Organization and functions of membrane proteins. Formation and release of transmembrane proteins in the cell The end product of the coupled work of transmembrane proteins

Cells. Binding to a signaling molecule (hormone or mediator) occurs on one side of the membrane, and the cellular response is formed on the other side of the membrane. Thus, they play a unique and important role in cell-to-cell communication and signal transduction.

Many transmembrane receptors consist of two or more subunits that act in combination and can dissociate upon binding to a ligand or change their conformation and move to the next stage of the activation cycle. They are often classified based on their molecular structure. The polypeptide chains of the simplest of these receptors cross the lipid bilayer only once, while many cross the lipid bilayer seven times (for example, G-protein-coupled receptors).

Structure

Extracellular domain

The extracellular domain is the region of the receptor that is located outside the cell or organoid. If the receptor polypeptide chain crosses the cell several times, then the outer domain may consist of several loops. The main function of the receptor is to recognize the hormone (although some receptors are also capable of responding to changes in membrane potential), and in many cases the hormone binds to this domain.

transmembrane domain

Some receptors are also protein channels. The transmembrane domain mainly consists of transmembrane α-helices. In some receptors, such as the nicotinic acetylcholine receptor, the transmembrane domain forms a membrane pore or ion channel. After activation of the extracellular domain (hormone binding), the channel can pass ions. In other receptors, after the binding of the hormone, the transmembrane domain changes its conformation, which has an intracellular effect.

intracellular domain

The intracellular, or cytoplasmic, domain interacts with the interior of the cell or organoid, relaying the received signal. There are two fundamentally different ways of such interaction:

• The intracellular domain binds to effector signaling proteins, which in turn transmit the signal along the signal chain to its destination.
• If the receptor is associated with an enzyme or itself has enzymatic activity, the intracellular domain activates the enzyme (or carries out an enzymatic reaction).

Classification

Most transmembrane receptors belong to one of three classes, distinguished by the main mechanism of signal transduction. Classify ionotropic and metabotropic transmembrane receptors. Ionotropic receptors, or receptors coupled to ion channels, are involved, for example, in the rapid transmission of synaptic signals between neurons and other target cells that can perceive electrical signals.

Metabotropic receptors transmit chemical signals. They are divided into two broad classes: G protein-coupled receptors and enzyme-coupled receptors.

G-protein coupled receptors are also called 7TM receptors (seven-transmembrane domain receptors, receptors with seven transmembrane domains). They are transmembrane proteins with an outer segment for ligand binding, a membrane segment, and a G protein-coupled cytosolic segment. They are divided into six classes based on the similarity of the structure and functions of the receptors, classes A-F(or 1-6), which, in turn, are divided into many families. This class includes sensory receptors and adrenoreceptors.

Like GPCRs, enzyme-coupled receptors are transmembrane proteins in which the ligand-binding domain is located outside the membrane. Unlike GPCRs, their cytosolic domain is not coupled to a G-protein, but itself has enzymatic activity or binds the enzyme directly. Usually, instead of seven segments, as in GPCRs, such receptors have only one transmembrane segment. These receptors may include the same signaling pathways as GPCRs. This class includes, for example, the insulin receptor.

There are six main classes of enzyme-coupled receptors:

• Receptor tyrosine kinases - can directly phosphorylate tyrosine residues, both their own and for a small set of intracellular signaling proteins.
• Tyrosine kinase-coupled receptors are not active enzymes themselves, but directly bind cytoplasmic tyrosine kinases for signal transduction.
• Receptor serine-threonine kinases - can directly phosphorylate serine or threonine residues, both their own and for the gene regulation proteins to which they bind.
• Histidine kinase-associated receptors activate a two-step signaling pathway in which the kinase phosphorylates its own histidine and immediately transfers phosphate to a second intracellular signaling protein.
• Receptor guanylate cyclases directly catalyze the production of cGMP molecules in the cytosol, which act as a small intracellular messenger in mechanisms very similar to cAMP.
• Receptor-like tyrosine phosphatases - remove phosphate groups from tyrosines of intracellular signaling proteins. They are called receptor-like because their mechanism of action as receptors remains unclear.

Regulation

In the cell, there are several ways to regulate the activity of transmembrane receptors, the most important ways are phosphorylation and internalization of receptors.

see also

Notes

Wikimedia Foundation. 2010 .

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Abstract on the topic:

Membrane proteins

Plan:

Introduction

• 1 Classification
  • 1.1 Topological classification
  • 1.2 Biochemical classification

Introduction

Alpha-helical transmembrane fragment of an integral protein.

To membrane proteins include proteins that are embedded in or associated with the cell membrane or the membrane of a cell organelle. About 25% of all proteins are membrane proteins.

1. Classification

Membrane proteins can be classified according to topological or biochemical principles. The topological classification is based on the location of the protein in relation to the lipid bilayer. Biochemical classification is based on the strength of the interaction of the protein with the membrane.

Different categories of polytopic proteins. Membrane binding via (1) single transmembrane alpha helix, (2) multiple transmembrane alpha helices, (3) beta sheet structure.

Various categories of integral monotopic proteins. Membrane binding by (1) amphipathic alpha helix parallel to the plane of the membrane, (2) hydrophobic loop, (3) covalently linked fatty acid residue, (4) electrostatic interaction (direct or calcium mediated).

1.1. Topological classification

In relation to the membrane, membrane proteins are divided into poly- and monotopic.

• Polytopic or transmembrane proteins completely penetrate the membrane and thus interact with both sides of the lipid bilayer. Typically, a transmembrane fragment of a protein is an alpha helix consisting of hydrophobic amino acids (possibly from 1 to 20 such fragments). Only in bacteria, as well as in mitochondria and chloroplasts, transmembrane fragments can be organized as a beta-sheet structure (from 8 to 22 turns of the polypeptide chain).
• Integral monotopic proteins permanently embedded in the lipid bilayer, but connected to the membrane only on one side without penetrating to the opposite side.

1.2. Biochemical classification

According to the biochemical classification, membrane proteins are divided into integral and peripheral.

• Integral membrane proteins are firmly embedded in the membrane and can only be removed from the lipid environment with the help of detergents or non-polar solvents. In relation to the lipid bilayer, integral proteins can be transmembrane polytopic or integral monotopic.
• Peripheral membrane proteins are monotopic proteins. They are either bound by weak bonds to the lipid membrane or are associated with integral proteins by hydrophobic, electrostatic, or other non-covalent forces. Thus, unlike integral proteins, they dissociate from the membrane when treated with an appropriate aqueous solution (eg, low or high pH, ​​high salt concentration, or chaotropic agent). This dissociation does not require the destruction of the membrane.

Membrane proteins can be incorporated into the membrane at the expense of fatty acid or prenyl residues or glycosylphosphatidylinositol attached to the protein during their post-translational modification.

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This abstract is based on an article from the Russian Wikipedia. Synchronization completed 07/14/11 05:26:08
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Principles of the structural organization of membrane proteins and methods for its prediction for transmembrane proteins

With high resolution managed to establish the structure of only one class of membrane proteins - the reaction center of bacteria, however, in this case, the position of the protein relative to the lipid bilayer is not unambiguously determined. The principles of its organization should be extended to other membrane proteins with caution. Some clarity can be introduced by using thermodynamic principles, as well as taking into account the fact that the bulk of experimental data is consistent with the assumption of a high content of α-helices in membrane proteins. Thermodynamic factors impose certain restrictions on what type of protein-lipid structures can be stable.

Membrane proteins are amphiphilic compounds

Any membrane proteins in direct contact with the hydrophobic core of the lipid bilayer must be amphiphilic. Those portions of the polypeptide that are exposed to solvent are likely to be enriched in polar and ionizable amino acid residues, while residues in contact with lylide hydrocarbon chains should be substantially non-polar. All this follows logically from the energy principles discussed in Sec. 2.3.1. Charged or polar amino acids can generally be located inside the bilayer, however, certain restrictions are imposed on this.

Let us consider three levels of amphiphilic structures in membrane proteins: primary, secondary and tertiary amphiphilicity.

1. Primary amphiphilic structures contain an extended stretch of predominantly non-polar amino acid residues, the length of which is sufficient to cross the bilayer. Such structures were found both in the reaction center and in bacteriorhodopsin. In these proteins, all elements penetrating the membrane are a-helical. The α-helical structure is preferred because it forms all hydrogen bonds in which the hydrogen atoms of the polypeptide backbone can participate. The alternative structure, which lacks one of the hydrogen bonds, is less stable by about 5 kcal/mol. All this allows us to suggest that the rotation of the polypeptide chain inside the membrane is unlikely. At the turning points, three to five amino acid residues would fail to form hydrogen bonds and this would destabilize the structure by about 15-20 kcal/mol. In globular, water-soluble proteins, the turning regions are located predominantly on the surface of the protein globule, where amide groups can form hydrogen bonds with water; Apparently, in the molecules of membrane proteins, the turns will also occur only in the regions exposed to water.

It is possible that the 3-layer can also form transmembrane elements having, for example, the shape of /J-cylinders, as in the case of porin. The requirements for the formation of hydrogen bonds by the hydrogen atoms of the polypeptide backbone in such structures can be satisfied, but only under the condition of interaction between individual ^-chains. How such a structure can be integrated into the membrane is not entirely clear, and the limitations imposed by the mechanisms of assembly of membrane proteins are generally unknown.

2. Secondary amphiphilic structures. In such structures, hydrophobic residues periodically occur along the chain, and when the polypeptide is folded into a certain secondary structure, they form a continuous surface. The periodicity of some elements of the secondary structure is indicated in Table. 1. Porins are an example of proteins in which secondary amphiphilic structures seem to play an important role. In them, polar and non-polar amino acid residues in each of the /3-chains alternate. All polar remnants are located on one side of the folded layer, lining a water-filled pore. Note that everything said about porin is hypothetical.

Table 1. Parameters of the secondary structure

Structure	Periodicity or number of residues per revolution	Distance between remnants, a	Radius or width, A
Uncurved
(Z-chain
Curved / Z-chain
Zu-Spiral
a-Spiral

a-helix, in which hydrophobic residues occur through every second or third monomeric unit, should have hydrophobic and polar surfaces. Such structures are often presented as a helical ring with side chains, as shown in Fig. Secondary amphiphilic structures can arise in the situations shown schematically in Fig. EVIL.

a. Surface-active protein segments; one side of the helix interacts with the hydrophobic region of the lipid bilayer, while the other side contacts the aqueous phase and the polar region of the bilayer. Amphiphilic a-helices are capable of forming many peptide hormones, as well as membrane-destroying peptides, such as mellitin.

b. transmembrane elements; the nonpolar surface of the helix faces the lipid phase, while the polar one lines the water channel penetrating the bilayer. This is a very common model, built mainly on the basis of the results of research on the nicotinic acetylcholine receptor, which functions as a chemically excitable channel. However, the conclusions based on experimental data that it is the amphiphilic helix that penetrates the membrane caused objections. Such a water-filled channel, as in a porin, can also form an amphiphilic 3-strand.

in. transmembrane elements; the non-polar part of the surface is in contact with lipids, and the polar groups are in contact with the polar groups of other transmembrane elements. It is this principle that underlies the "inverted" structures, which is supposedly bacteriorhodopsin. Polar interactions between amphiphilic helices could in principle stabilize interactions between subunits in oligomeric proteins.

3. Tertiary amphiphilic structures. Their existence can only be hypothesized. Their hydrophobic surface should be formed at the level of the tertiary structure of the residues located in the most different parts of the polypeptide chain. Such structures may be characteristic of proteins that bind to the bilayer, but do not have well-defined hydrophobic domains, determined by any of the above criteria. A possible example of this kind is a-lactalbumin.

Ionizable amino acid residues in transmembrane segments

Many models of membrane proteins suggest that their transmembrane segments contain ionizable residues. These residues likely play an important functional and/or structural role. In some cases, this role has been unequivocally established: 1) lysine residues in bacteriorhodopsin and rhodopsin form Schiff bases with a retinal prosthetic group, which is necessary for light excitation of the molecule; 2) histidine residues in the polypeptides of the bacterial reaction center are involved in binding to photosynthetic pigments; 3) charged residues in lactose permease from E. coli are involved in the transport functions of this protein; it is possible that these residues form a network of hydrogen bonds within the protein molecule.

The transfer of charged groups from water to a medium with a low permittivity inside the membrane is energetically very unfavorable, and these groups must be stabilized in some way. It has been repeatedly assumed that the formation of ion pairs is sufficient for stabilization, and this principle was used to construct a three-dimensional model of bacteriorhodopsin. However, calculations have shown that the free energy of ion pair transfer from water to a medium with a low permittivity is also very high. Further stabilization requires additional polar interactions, possibly involving other polar groups or via hydrogen bonds.

In principle, even a single charged group inside the membrane can be stabilized through interactions with polar groups and with the participation of hydrogen bonds that effectively delocalize the charge. There are several examples of isolated, desolvated ions stabilized by interactions in water-soluble proteins. Similar principles seem to operate in the case of charged residues of transmembrane segments of integral proteins.

However, it seems more likely that ionizable amino acids are neutralized within the membrane by protonation or deprotonation. The neutralization free energy of charged amino acids has been estimated to be approximately 10-17 kcal/mol. In the absence of specific conditions for polar interactions that stabilize the charged residue in the transmembrane segment, it is likely to be neutralized.

Charged amino acids in segments exposed to the aquatic environment

As we have already said, the charged residues are distributed asymmetrically between the two sides of the bacterial reaction center. This asymmetry is also characteristic of some other internal membrane proteins of bacteria. Thus, the main residues of Lys and Arg are four times more common in those regions connecting transmembrane elements that are located on the inner side of the membrane, and not on the outer. For the acid residues Asp and Glu, no such trend is revealed. It is possible that this asymmetry is related to the assembly mechanism of the membrane protein, but how exactly is unclear. Moreover, it is not known whether this observation can be generalized and whether it has any predictive value.

In globular, water-soluble proteins, proline residues are rarely located in the middle part of the a-helix. According to the results of studies of 58 proteins containing 331 α-helices, 30 such cases were identified. In half of them, proline was located at the sites of damage to the helix, and in the remaining cases, it was located in the area of ​​curvature or irregularity of the structure.

At the same time, in bacteriorhodopsin, proline residues are located in the middle part of three of the seven transmembrane helices, and in rhodopsin - in five of the seven such spirals. A similar trend was also found for other transmembrane segments of integral proteins, especially transport ones. The significance of this phenomenon is unknown. It should be noted, however, that due to the presence of a cyclic side chain, proline does not form hydrogen bonds with residues located on the previous turn of the a-helix. This may contribute to the formation of structures in which the hydrogen bond is formed due to a specific interaction with a residue located in another membrane-permeating region. Such a polar interaction within the bilayer could stabilize the three-dimensional structure of membrane proteins.

Methods for identifying primary amphiphilic structures

Unambiguous structural information about membrane proteins has been obtained only in a few cases, but researchers have extensive amino acid sequence data based on DNA sequencing results. To identify transmembrane α-helices, presumably 20 residues long and consisting predominantly of hydrophobic amino acids, several amino acid sequence analysis methods have been developed. Each of them is based on the arrangement of amino acids in a row in accordance with a certain parameter that reflects the probability of finding this residue in the transmembrane segment.

There are two types of scales. In one instance, amino acids are classified by their relative polarity or "hydrophobicity". These scales are of a thermodynamic nature and are based on the magnitude of the change in free energy during the transfer of an amino acid from an aqueous solution into a hydrocarbon medium. However, the number of methods for quantifying the hydrophobicity of amino acids is very large, and they do not agree with each other in everything. Often, data is used that relates simultaneously to more than one physical characteristic. An example of this kind is the "hydropathy" scale of Kite and Doolittle, based on data on hydrophobicity, measured by hydration potential, as well as the probability of finding residues inside the globule.

The Goldman, Engelman, and Steitz scale is based on a quantitative assessment of the free energy of the transfer of a-helices from an aqueous medium into the membrane. On fig. the Kite-Doolittle scale is compared with the Goldman-Engelman-Steitz scale.

According to Engelman et al., the change in free energy upon the introduction of a polyanionic a-helix 20 residues long into the membrane is 30 kcal/mol. The calculation is based on an estimate of the surface area of ​​the helix exposed to the solvent. The contribution to the energy of each side group was evaluated taking into account the surface area exposed to the aqueous medium inside the helix. The free energy of transfer into the bilayer of polar groups was also taken into account. For example, it was assumed that glutamine would be protonated upon transfer to the bilayer and the free energy of this process would be 10.8 kcal/mol. Likewise, the transfer of hydroxyls would "cost" approximately 4.0 kcal/mol.

All of the above shows how the gain in interaction energy upon transfer of the α-helix into the bilayer can be used to "draw" polar side groups into the bilayer. For example, one arginine residue can be incorporated into a bilayer as part of a nonpolar transmembrane helix if it is deprotonated; this requires 16.7 kcal/mol at pH 7.0. The total free energy of transfer of the a-helix will still remain negative. However, the situation will change if two arginine residues need to be inserted into the bilayer or if arginine is positively charged. Of course, polar residues can be stabilized within the bilayer due to specific interactions, but it is very difficult to take this into account in calculations. For example, serine, cysteine, and threonine side groups can form hydrogen bonds to the polypeptide backbone, and acidic and basic residues can form ion pairs; the appearance of such pairs is possible if these residues are located four or five monomeric units apart.

The second type of scale, which is used to classify amino acids, is based on data on the frequency with which amino acids actually occur in membrane-spanning segments.

This empirically takes into account hydrophobicity, as well as many other factors that cannot be quantified as hydrophobicity. The disadvantage of this semi-empirical approach is the lack of accurate data on the boundaries of the transmembrane regions. However, such scales can be just as useful as scales based on thermodynamic parameters. Examples include Kuhn and Leigh's "tendency" scale to the membrane, or Rao and Argos' "helix immersion into the membrane" scale. The four most hydrophobic residues on the Goldman-Engelmann-Steitz scale are also the four residues with the highest parameter value on the Rao and Argos scale.

On fig. profiles of three different membrane proteins obtained using different scales are presented. When constructing these profiles, the average values ​​of the numbers on the scales assigned to each amino acid within the selected “window” are taken into account; this average is plotted relative to the number of the residue in the polypeptide. For example, if the "window" is 19 residues, the value assigned to position 40 would be the average on the scale for all amino acids from 31 to 49 inclusive. The value assigned to position 41 will be the average of residues 32 to 50, and so on. The peaks on the profile correspond to hydrophobic regions or those regions that are more likely to form transmembrane helices. To build a profile, the size of the window is important; most of the curves in Fig. were built with a window size of 19 residuals.

Let's try to interpret the constructed profiles. According to the Goldman-Engelman-Steitz scale, peaks at values ​​close to zero correspond to transmembrane helices. The value of 1.25 on the Kite-Doolittle scale is the smallest value corresponding to the known transmembrane helix in the L subunit of the reaction center R . viridis . In all three cases shown in Fig. 3.12, the profiles for the reaction center subunits are similar.

On fig. two profiles for cytochrome P450 from microsomes are shown. This protein was chosen because its primary structure data suggest that it has eight transmembrane helices. However, the available experimental data indicate the existence of only one N- koh- end anchor in the membrane. Both the Kite-Doolittle profile and the Goldman-Engelman-Steitz profile reveal the N-terminal region, but they also indicate the presence of one or more additional transmembrane segments, which is not true. Note that many of the constructed models of membrane proteins, which are based only on amino acid sequence data, may be incorrect.

On fig. three profiles for bacteriorhodopsin are given. Despite their similarity, differences are seen in the shape of the peaks corresponding to the seven transmembrane segments. The Goldman-Engelman-on-Steitz algorithm does not take into account the stabilizing effect associated with the formation of an ion pair from closely spaced charged residues within one helix. With this factor in mind, the division between the last two spirals becomes clearer.

One of the problems faced by the application of all the algorithms described above is to exclude hydrophobic segments in known globular proteins that are not transmembrane, but are located inside the protein. However, when we are looking for sufficiently long sections, this problem does not arise.

Note that the algorithms used to detect a-helical structures in soluble globular proteins, such as the Chow-Fasman algorithm, are not suitable for detecting transmembrane elements. These algorithms are not applicable to describe the structure of non-globular regions, such as segments located inside the bilayer.

Algorithms designed to identify transmembrane regions cannot be used in the case of segments that are secondary amphiphilic structures or cross the membrane in the form of a /3 layer. In the first case, this region is excluded from consideration due to the presence of polar residues in it, and in the second, the transmembrane segment is too short, since only 10–12 amino acid residues in the /3 structure are needed to cross the bilayer. Some algorithms were designed to detect ^-turns rather than the transmembrane elements themselves. Although this avoids some of the problems associated with isolating different classes of transmembrane elements, it is not clear how acceptable they will be with their wider application.

Methods for identifying secondary amphiphilic structures

Several approaches have been developed to detect secondary amphiphilicity or asymmetry in the distribution of hydrophobic residues in polypeptide chain segments. Quite often, a-helices and /3-layers in globular proteins are characterized by periodicity in the distribution of hydrophobic residues. The use of a spiral ring as a qualitative indicator is not always justified; more quantitative approaches are needed. The main one is the determination of the periodicity in the distribution of hydrophobic residues using Fourier transform methods. An example is the hydrophobic moment.

1. hydrophobic moment. This parameter was proposed by Eisenberg et al. It is defined as

and represents a certain vector sum of the hydrophobicity of residues in a segment of N elements. The hydrophobicity of each residue is represented as a vector, which is characterized by the angle , formed by the side chain and the axis of the polypeptide backbone. For a-helix 6 = 100°. On fig. 3.9 B The "vectors" of hydrophobicity are presented in projection onto the plane of the spiral ring, and the hydrophobic moment is equal to their vector sum. The hydrophilic residue is represented by a vector with negative directionality. For a random sequence, the value qi by virtue of random distribution there will be very few hydrophobic residues. At the same time, in the mellitin peptide, hydrophobic residues are located on one side of the structure, and polar ones, on the other. The numerical value of the hydrophobic moment is assigned to the amino acid located in the center of the analyzed segment. Therefore, one can “scan” the sequence and assign average hydrophobicity to each position, and also find

Eisenberg et al. analyzed 11-residue-long segments from many proteins and peptides, determining the hydrophobic moment

and average hydrophobicity for each of the studied segments. Polypeptide segments of globular proteins are characterized by low values ​​of both and qi - Transmembrane elements of a hydrophobic nature have high but low pH values, being mostly non-polar. Peptides and regions of proteins related to "surfactant" have high values tsn due to strong asymmetry in the distribution of polar and non-polar residues. Using this algorithm, some segments of surfactant proteins were identified, such as diphtheria toxin and pyruvate oxidase regions from E. coli.

The hydrophobic moment serves as a quantitative measure of the periodicity in the distribution of hydrophobic residues in different regions of the polypeptide. Choice 6 plays an important role here. The hydrophobic moment is essentially one of the parameters of the Fourier transform of the hydrophobicity function. More general methods, described below, allow us to analyze all the Fourier components and identify any possible periodicity.

2. Periodicity of the sequence. Many methods have been developed for identifying regions of protein molecules that are characterized by periodic changes in hydrophobicity along the chain. All of them involve the Fourier transform of a function dependent on the hydrophobicity of the amino acid residues along the polypeptide. The presence of a peak with a period of 3.6 indicates that a hydrophobic residue in this segment of the analyzed polypeptide occurs on average every 3.6 residues. This means that the segment is an α-helix, on one side of which there are predominantly hydrophobic residues. This method has been used to identify amphiphilic regions in some transport and channel proteins; examples include the acetylcholine receptor, the sodium channel, the glucose transporter, the mitochondrial uncoupler protein, and the erythrocyte band 3 protein, which is an anion transporter. However, there is no clear indication that these putative amphiphilic helices are transmembrane.

These methods have also been used to analyze membrane surface interacting peptides and apolipoproteins.

Peptides - models of membrane proteins

Peptides began to be used to study protein-lipid interactions many years ago. In most cases, these were natural membrane-active peptides, primarily gramicidin A, alamethicin, and mellitin. Currently, synthetic peptides are more often used as model systems. In this case, it is necessary to remember two points: 1) when binding the peptide to the membrane, both primary and secondary amphiphilicities are essential; 2) peptides often have polymorphism, i.e. the ability to change conformation depending on the environment. Not; It is possible that in the future with the help of synthetic peptides it will be possible to study protein-lipid interactions in detail, but so far we are still very far from this.

The formation of transmembrane proteins should include the stages of recognition of transmembrane domains and their integration into the lipid bilayer.

Transmembrane domains exit the translocon laterally across the protein-lipid interface

Positioning on the translocation channel and the beginning transport of secretory and transmembrane proteins happen the same way. However, the translocation of membrane proteins must be combined with their integration or insertion into the ER lipid bilayer. Integration occurs at the moment when the transmembrane domains are recognized by the translocon, then their translocation into the ER lumen stops, and they begin to be transferred from the channel to the lipid bilayer in the lateral direction. In this way, many various types transmembrane proteins, including those that span the membrane multiple times.

first step on the path of protein integration into the membrane is the recognition of transmembrane domains by the translocon. These domains extend for about twenty hydrophobic amino acids. Due to their hydrophobic nature, some transmembrane domains are recognized by SRPs as signal sequences. These so-called signal anchor sequences first position the newly formed protein on the ER, and then are directed to the channel as normal signal sequences.

However signal anchor sequences are not cleaved from the protein, but are integrated into the membrane. As shown in the figure below, unlike the signal anchor sequence, most transmembrane domains are recognized by the translocon as soon as they leave the ribosome, after addressing is completed by the normal N-terminal signal sequence. The information that the transmembrane domain has already been synthesized must be transmitted to the translocon by a different route than from SRP.

Signal anchor sequences are transferred directly from the SRP to the translocon,
however, recognition of internal transmembrane domains must occur as they are released from the ribosome.

Simplest sign, indicating that the transmembrane domain is located in the translocon, this is the hydrophobicity of the domain itself. Due to the peculiarity of the structure, the translocation channel exhibits this hydrophobicity. As shown in fig. 3.21, the structure of the translocon suggests that the channel is able to open like a clam shell, allowing the transmembrane domain to simultaneously contact the channel and the lipid bilayer. Apparently, the signal sequences and transmembrane domains bind to the Sec61a protein located on the side of the opening valves, and this binding then causes the lateral opening of the channel.

This scheme has been proposed based on experimental data, according to which the transmembrane domains in the channel are in contact with Sec61a and . As a result, although the translocon contains an aqueous channel, there are enough hydrophobic channels in the membrane, in which translocated polypeptides can enter the lipid environment. It should be expected that regions containing polar amino acids should move through the channel without stopping, while hydrophobic domains, due to strong interaction with lipids, will remain associated with the side walls of the channel, preventing translocation.

In some cases translocon may otherwise identify the transmembrane domain. For example, sometimes during the synthesis of a transmembrane domain, changes in the interaction between the ribosome and the translocon occur before the domain has left the ribosome. These changes serve as a signal for the translocon that a transmembrane domain is about to appear. How the transmembrane domain induces changes in the ribosome and transmits them to the translocon remains unclear. Sometimes recognition also requires polar elements of the newly formed chain adjacent to the transmembrane domain. This suggests that, at least in some cases, the recognition process must involve more than just a hydrophobic interaction between the domain and the lipid-channel environment.

The border of the channel and the lipid layer seems to serve as a pathway for transmembrane domains to exit the channel after they are recognized. However, the mechanism of domain exit from the translocon somewhat varies from substrate to substrate. Some domains leave the translocon almost immediately after being recognized in the channel. In these cases, the transmembrane domain first contacts Sec61a and lipids, and then only lipids; it is assumed that the domain has already penetrated into the lipid bilayer.

For integration of such domains of other proteins, with the exception of the Sec61 complex, is not required. Other transmembrane domains integrate more slowly and, after recognition, do not leave the translocon for a long time, sometimes even remaining there until the end of translation. As they exit the channel into the bilayer, these transmembrane domains come into contact with the TRAM protein, although its further role remains unclear.

Degree of hydrophobicity determines in part whether the transmembrane domain integrates immediately or occurs at a later stage in protein synthesis. More hydrophobic domains may move faster into the lipid bilayer, but less hydrophobic ones may remain at the boundary and require additional transport factors. It is possible that TRAM and other proteins serve as chaperones for some transmembrane domains. They promote the integration of such domains, the hydrophobicity of which is insufficient for movement.

Obviously, at least group of transmembrane proteins may represent multiple forms containing a specific domain, which in some cases is integrated into the membrane, while in others it remains unrecognized. Proteins such as TRAM can determine under what conditions such substitutions will be integrated.

The translocon is depicted as a cylinder,
which opens and closes in two ways,
allowing the movement of the newly formed chain through the pore and the promotion of the transmembrane domain into the membrane.
A gap in the wall of the translocation channel allows proteins to enter the lipid bilayer,
and to recognize and integrate transmembrane domains.
Since the domains are hydrophobic, they prefer the lipid environment and migrate out of the channel into the lipid bilayer.

If the main role of lipids in the composition of membranes is to stabilize the bilayer, then proteins are responsible for the functional activity of membranes. Some of them provide transport of certain molecules and ions, others are enzymes, others are involved in the binding of the cytoskeleton to the extracellular matrix or serve as receptors for hormones, mediators,

eicosanoids, lipoproteins, nitric oxide (N0). Proteins account for 30 to 70% of the mass of membranes. Proteins determine the features of the functioning of each membrane.

Structural features

and localization of proteins in membranes

Membrane proteins in contact with the hydrophobic part of the lipid bilayer must be amphiphilic. Those parts of the protein that interact with hydrocarbon chains of fatty acids contain predominantly non-polar amino acids. The regions of the protein located in the region of the polar "heads" are enriched in hydrophilic amino acid residues.

Localization of proteins in membranes. Transmembrane proteins, for example: 1 - glycophorin A; 2 - adrenaline receptor. Surface proteins: 3 - proteins associated with integral proteins, for example, the enzyme succinate dehydrogenase; 4 - proteins attached to the polar "heads" of the lipid layer, for example, protein kinae C; 5 - proteins "anchored" in the membrane using a short hydrophobic terminal domain, for example, cytochromes b 5; 6 - "anchored" proteins covalently connected to the membrane pipid (for example, the enzyme alkaline phosphatase).

Membrane proteins differ in their position in the membrane. They can penetrate deeply into the lipid bilayer or even permeate it - integral proteins, or attach to the membrane in different ways - surface proteins.

Surface proteins

Surface proteins often attach to the membrane by interacting with integral

proteins or surface areas of the lipid layer.

Proteins that form complexes with integral membrane proteins

A number of digestive enzymes involved in the hydrolysis of starch and proteins are attached to the integral proteins of the intestinal microvilli membranes.

Examples of such complexes are sucrase-isomaltase and maltase-glycoamylase.

Proteins Associated with Polar Heads of Membrane Lipids

Polar or charged domains of a protein molecule can interact with the polar "heads" of lipids, forming ionic and hydrogen bonds. In addition, many proteins soluble in the cytosol can, under certain conditions, bind to the membrane surface for a short time. Sometimes protein binding is a necessary condition for the manifestation of enzymatic activity. Such proteins, for example, include protein kinase C, blood coagulation factors.

Anchoring with a membrane "anchor"

The "anchor" can be a non-polar domain of the protein, built from amino acids with hydro-

phobic radicals. An example of such a protein is cytochrome b 5 of the ER membrane. This protein is involved in redox reactions as an electron carrier.

The role of the membrane "anchor" can also be performed by a fatty acid residue covalently bound to the protein (myristic - C 14 or palmitic - C 16). Proteins associated with fatty acids are localized mainly on the inner surface of the plasma membrane. Myristic acid adds to the N-terminal glycine to form an amide bond. Palmitic acid forms a thioether bond with cysteine ​​or an ester bond with serine and threonine residues.

A small group of proteins can interact with the outer surface of the cell using a phosphatidylinositolglycan covalently attached to the C-terminus of the protein. This "anchor" is often the only link between the protein and the membrane, therefore, under the action of phospholipase C, this protein is separated from the membrane.

Transmembrane (integral) proteins

Some of the transmembrane proteins penetrate the membrane once (glycophorin), others have several sections (domains) that successively cross the bilayer.

The transmembrane domains penetrating the bilayer have an α-helix conformation. Polar amino acid residues face inside the globule, while non-polar ones contact with membrane lipids. Such proteins are called "inverted" compared to water-soluble proteins, in which most of the hydrophobic amino acid residues are hidden inside, and the hydrophilic ones are located on the surface.

The charged amino acid radicals in these domains are uncharged and protonated (-COOH) or deprotonated (-NH 2).

Glycosylated proteins

Surface proteins or domains of integral proteins located on the outer surface of all membranes are almost always glycosylated. Oligosaccharide Residues can be attached through the amide group of asparagine or the hydroxyl groups of serine and threonine.

Oligosaccharide residues protect the protein from proteolysis and are involved in ligand recognition or adhesion.

Lateral diffusion of proteins

Some membrane proteins move along the bilayer (lateral diffusion) or rotate around an axis, perpendicular to its surface.

Lateral diffusion of integral proteins in the membrane is limited, this is due to their large size, interaction with other membrane proteins, elements of the cytoskeleton or extracellular matrix.

Membrane proteins do not move from one side of the membrane to the other (“flip-flop” jumps), like phospholipids.