Protein-RNA interactions: structural analysis and functional classes

A data set of 89 protein-RNA complexes has been extracted from the Protein Data Bank, and the nucleic acid recognition sites characterized through direct contacts, accessible surface area, and secondary structure motifs. The differences between RNA recognition sites that bind to RNAs in functional classes has also been analyzed. Analysis of the complete data set revealed that van der Waals interactions are more numerous than hydrogen bonds and the contacts made to the nucleic acid backbone occur more frequently than specific contacts to nucleotide bases. Of the base-specific contacts that were observed, contacts to guanine and adenine occurred most frequently. The most favored amino acid-nucleotide pairings observed were lysine-phosphate, tyrosine-uracil, arginine-phosphate, phenylalanine-adenine and tryptophan-guanine. The amino acid propensities showed that positively charged and polar residues were favored as expected, but also so were tryptophan and glycine. The propensities calculated for the functional classes showed trends similar to those observed for the complete data set. However, the analysis of hydrogen bond and van der Waal contacts showed that in general proteins complexed with messenger RNA, transfer RNA and viral RNA have more base specific contacts and less backbone contacts than expected, while proteins complexed with ribosomal RNA have less base-specific contacts than the expected. Hence, whilst the types of amino acids involved in the interfaces are similar, the distribution of specific contacts is dependent upon the functional class of the RNA bound.     

Dissecting the protein-RNA interface: the role of protein surface shapes and RNA secondary structures in protein-RNA recognition

Protein-RNA interactions are essential for many biological processes. However, the structural mechanisms underlying these interactions are not fully understood. Here, we analyzed the protein surface shape (dented, intermediate or protruded) and the RNA base pairing properties (paired or unpaired nucleotides) at the interfaces of 91 protein-RNA complexes derived from the Protein Data Bank. Dented protein surfaces prefer unpaired nucleotides to paired ones at the interface, and hydrogen bonds frequently occur between the protein backbone and RNA bases. In contrast, protruded protein surfaces do not show such a preference, rather, electrostatic interactions initiate the formation of hydrogen bonds between positively charged amino acids and RNA phosphate groups. Interestingly, in many protein-RNA complexes that interact via an RNA loop, an aspartic acid is favored at the interface. Moreover, in most of these complexes, nucleotide bases in the RNA loop are flipped out and form hydrogen bonds with the protein, which suggests that aspartic acid is important for RNA loop recognition through a base-flipping process. This study provides fundamental insights into the role of the shape of the protein surface and RNA secondary structures in mediating protein-RNA interactions.     

Hydration of protein-protein interfaces

We present an analysis of the water molecules immobilized at the protein-protein interfaces of 115 homodimeric proteins and 46 protein-protein complexes, and compare them with 173 large crystal packing interfaces representing nonspecific interactions. With an average of 15 waters per 1000 A2 of interface area, the crystal packing interfaces are more hydrated than the specific interfaces of homodimers and complexes, which have 10-11 waters per 1000 A2, reflecting the more hydrophilic composition of crystal packing interfaces. Very different patterns of hydration are observed: Water molecules may form a ring around interfaces that remain "dry," or they may permeate "wet" interfaces. A majority of the specific interfaces are dry and most of the crystal packing interfaces are wet, but counterexamples exist in both categories. Water molecules at interfaces form hydrogen bonds with protein groups, with a preference for the main-chain carbonyl and the charged side-chains of Glu, Asp, and Arg. These interactions are essentially the same in specific and nonspecific interfaces, and very similar to those observed elsewhere on the protein surface. Water-mediated polar interactions are as abundant at the interfaces as direct protein-protein hydrogen bonds, and they may contribute to the stability of the assembly.     

Extent and nature of contacts between protein molecules in crystal lattices and between subunits of protein oligomers

A survey was compiled of several characteristics of the intersubunit contacts in 58 oligomeric proteins, and of the intermolecular contacts in the lattice for 223 protein crystal structures. The total number of atoms in contact and the secondary structure elements involved are similar in the two types of interfaces. Crystal contact patches are frequently smaller than patches involved in oligomer interfaces. Crystal contacts result from more numerous interactions by polar residues, compared with a tendency toward nonpolar amino acids at oligomer interfaces. Arginine is the only amino acid prominent in both types of interfaces. Potentials of mean force for residue–residue contacts at both crystal and oligomer interfaces were derived from comparison of the number of observed residue–residue interactions with the number expected by mass action. They show that hydrophobic interactions at oligomer interfaces favor aromatic amino acids and methionine over aliphatic amino acids; and that crystal contacts form in such a way as to avoid inclusion of hydrophobic interactions. They also suggest that complex salt bridges with certain amino acid compositions might be important in oligomer formation. For a protein that is recalcitrant to crystallization, substitution of lysine residues with arginine or glutamine is a recommended strategy. Proteins 28:494–514, 1997. © 1997 Wiley-Liss, Inc.     

Protein-nucleic acid recognition: statistical analysis of atomic interactions and influence of DNA structure

We analyzed structural features of 11,038 direct atomic contacts (either electrostatic, H-bonds, hydrophobic, or other van der Waals interactions) extracted from 139 protein-DNA and 49 protein-RNA nonhomologous complexes from the Protein Data Bank (PDB). Globally, H-bonds are the most frequent interactions (approximately 50%), followed by van der Waals, hydrophobic, and electrostatic interactions. From the protein viewpoint, hydrophilic amino acids are over-represented in the interaction databases: Positively charged amino acids mainly contact nucleic acid phosphate groups but can also interact with base edges. From the nucleotide point of view, DNA and RNA behave differently: Most protein-DNA interactions involve phosphate atoms, while protein-RNA interactions involve more frequently base edge and ribose atoms. The increased participation of DNA phosphate involves H-bonds rather than salt bridges. A statistical analysis was performed to find the occurrence of amino acid-nucleotide pairs most different from chance. These pairs were analyzed individually. Finally, we studied the conformation of DNA in the interaction sites. Despite the prevalence of B-DNA in the database, our results suggest that A-DNA is favored in the interaction sites.     

Interaction geometry involving planar groups in protein-protein interfaces

The geometry of interactions of planar residues is nonrandom in protein tertiary structures and gives rise to conventional, as well as nonconventional (X--H...pi, X--H...O, where X = C, N, or O) hydrogen bonds. Whether a similar geometry is maintained when the interaction is across the protein-protein interface is addressed here. The relative geometries of interactions involving planar residues, and the percentage of contacts giving rise to different types of hydrogen bonds are quite similar in protein structures and the biological interfaces formed by protein chains in homodimers and protein-protein heterocomplexes--thus pointing to the similarity of chemical interactions that occurs during protein folding and binding. However, the percentage is considerably smaller in the nonspecific and nonphysiological interfaces that are formed in crystal lattices of monomeric proteins. The C--H...O interaction linking the aromatic and the peptide groups is quite common in protein structures as well as the three types of interfaces. However, as the interfaces formed by crystal contacts are depleted in aromatic residues, the weaker hydrogen bond interactions would contribute less toward their stability.     

Discovering the interaction propensities of amino acids and nucleotides from protein-RNA complexes

With the availability of many genome sequences, the mining of biological data is attracting much attention, most of it limited to the sequences of macromolecules. Sequence data are easy to analyze as they can be treated as strings of characters, whereas the structure of a macromolecule is much more complex. We developed a set of algorithms to analyze the structures of protein-RNA complexes at the atomic level and used them to analyze protein-RNA interactions using structural data on 51 protein-RNA complexes. The analysis revealed, among other things, that: (1) polar and charged amino acids have a strong tendency to interact with nucleotides, (2) arginine and asparagine tend to hydrogen bond with uracil, and (3) histidine favors uracil in water-mediated bonding with RNA. We analyzed a large set of structural data of protein-RNA complexes involving water-mediated hydrogen bonds as well as direct hydrogen bonds. The interaction patterns discovered from the analysis provide useful information for predicting the structure of RNA that binds proteins, and of proteins that bind RNA.     

Intramolecular surface contacts contain information about protein-protein interface regions

MOTIVATION: Some amino acids clearly show preferences over others in protein-protein interfaces. These preferences, or so-called interface propensities can be used for a priori interface prediction. We investigated whether the prediction accuracy could be improved by considering not single but pairs of residues in an interface. Here we present the first systematic analysis of intramolecular surface contacts in interface prediction. RESULTS: We show that preferences do exist for contacts within and around an interface region within one molecule: specific pairs of amino acids are more often occurring than others. Using intramolecular contact propensities in a blind test, higher average scores were assigned to interface residues than to non-interface residues. This effect persisted as small but significant when the contact propensities were corrected to eliminate the influence of single amino acid interface propensity. This indicates that intramolecular contact propensities may replace interface propensities in protein-protein interface prediction. AVAILABILITY: The source code is available on request from the authors.     

Correlation of sequence and tertiary structure in globular proteins

The x-ray crystal structures of 19 selected proteins are examined empirically for correlations between the amino acid sequence and long-range, tertiary conformation. There is clear evidence for preferential associations of certain types of amino acids, particularly among the hydrophobic aliphatic, aromatic, and cysteine residues. However, the likelihoods of forming these residue-pair contacts are all less than 12%, so packing and geometric requirements must often take precedent over energetic considerations. The prediction of long-range contacts is not substantially improved by taking into account the sequentially previous residues. The analysis of atom–atom contacts shows a similar lack of predictive ability, but the results show that a good approximation to the interresidue energy function must include different types of interactions at two or three different sites on some amino acids. Backbone–backbone long-range interactions are relatively rare and nonspecific, whereas some “polar” side chains form hydrogen bonds from the polar groups while occasionally forming hydrophobic contacts with the remainder of the chain.     

The role of RNA sequence and structure in RNA--protein interactions

We investigate the sequence and structural properties of RNA-protein interaction sites in 211 RNA-protein chain pairs, the largest set of RNA-protein complexes analyzed to date. Statistical analysis confirms and extends earlier analyses made on smaller data sets. There are 24.6% of hydrogen bonds between RNA and protein that are nucleobase specific, indicating the importance of both nucleobase-specific and -nonspecific interactions. While there is no significant difference between RNA base frequencies in protein-binding and non-binding regions, distinct preferences for RNA bases, RNA structural states, protein residues, and protein secondary structure emerge when nucleobase-specific and -nonspecific interactions are considered separately. Guanine nucleobase and unpaired RNA structural states are significantly preferred in nucleobase-specific interactions; however, nonspecific interactions disfavor guanine, while still favoring unpaired RNA structural states. The opposite preferences of nucleobase-specific and -nonspecific interactions for guanine may explain discrepancies between earlier studies with regard to base preferences in RNA-protein interaction regions. Preferences for amino acid residues differ significantly between nucleobase-specific and -nonspecific interactions, with nonspecific interactions showing the expected bias towards positively charged residues. Irregular protein structures are strongly favored in interactions with the protein backbone, whereas there is little preference for specific protein secondary structure in either nucleobase-specific interaction or -nonspecific interaction. Overall, this study shows strong preferences for both RNA bases and RNA structural states in protein-RNA interactions, indicating their mutual importance in protein recognition.     

Non-covalent interactions across subunit interfaces in Sm proteins

The distinguishing property of Sm protein associations is their high stability. In order to understand this property, we analyzed the interface non-covalent interactions and compared the properties of the Sm protein interfaces with those of a test set, Binding Interface Database (BID). The comparison revealed that the main differences between interfaces of Sm proteins and those of the BID set are the content of charged residues, hydrogen bonds, salt bridges, and conservation scores of interface residues. In Sm proteins, the interfaces have more hydrophobic and fewer charged residues than the surface, which is also the case for the BID test set and other proteins. However, in the interfaces, the content of charged residues in Sm proteins (26%) is substantially larger than that in the BID set (22%). Both interfaces of Sm proteins and of test set have a similar number of hydrophobic interactions per 100 Å². The interfaces of Sm proteins have substantially more hydrogen bonds than the interfaces in test set. The results show clearly that the interfaces of Sm proteins form more salt bridges compared with test set. On average, there are about 16 salt bridges per interface. The high conservation score of amino acids that are involved in non-covalent interactions in protein interfaces is an additional strong argument for their importance. The overriding conclusion from this study is that the non-covalent interactions in Sm protein interfaces considerably contribute to stability of higher order structures.     

Landscape of π–π and sugar–π contacts in DNA–protein interactions

There were 1765 contacts identified between DNA nucleobases or deoxyribose and cyclic (W, H, F, Y) or acyclic (R, E, D) amino acids in 672 X-ray structures of DNA–protein complexes. In this first study to compare π-interactions between the cyclic and acyclic amino acids, visual inspection was used to categorize amino acid interactions as nucleobase π–π (according to biological edge) or deoxyribose sugar–π (according to sugar edge). Overall, 54% of contacts are nucleobase π–π interactions, which involve all amino acids, but are more common for Y, F, and R, and involve all DNA nucleobases with similar frequencies. Among binding arrangements, cyclic amino acids prefer more planar (stacked) π-systems than the acyclic counterparts. Although sugar–π interactions were only previously identified with the cyclic amino acids and were found to be less common (38%) than nucleobase–cyclic amino acid contacts, sugar–π interactions are more common than nucleobase π–π contacts for the acyclic series (61% of contacts). Similar to DNA–protein π–π interactions, sugar–π contacts most frequently involve Y and R, although all amino acids adopt many binding orientations relative to deoxyribose. These DNA–protein π-interactions stabilize biological systems, by up to approximately ?40 kJ mol^?1 for neutral nucleobase or sugar–amino acid interactions, but up to approximately ?95 kJ mol^?1 for positively or negatively charged contacts. The high frequency and strength, despite variation in structure and composition, of these π-interactions point to an important function in biological systems.     

A knowledge-based potential function predicts the specificity and relative binding energy of RNA-binding proteins

RNA-protein interactions are fundamental to gene expression. Thus, the molecular basis for the sequence dependence of protein-RNA recognition has been extensively studied experimentally. However, there have been very few computational studies of this problem, and no sustained attempt has been made towards using computational methods to predict or alter the sequence-specificity of these proteins. In the present study, we provide a distance-dependent statistical potential function derived from our previous work on protein-DNA interactions. This potential function discriminates native structures from decoys, successfully predicts the native sequences recognized by sequence-specific RNA-binding proteins, and recapitulates experimentally determined relative changes in binding energy due to mutations of individual amino acids at protein-RNA interfaces. Thus, this work demonstrates that statistical models allow the quantitative analysis of protein-RNA recognition based on their structure and can be applied to modeling protein-RNA interfaces for prediction and design purposes.     

i‐Patch: Interprotein contact prediction using local network information

Biological processes are commonly controlled by precise protein‐protein interactions. These connections rely on specific amino acids at the binding interfaces. Here we predict the binding residues of such interprotein complexes. We have developed a suite of methods, i‐Patch, which predict the interprotein contact sites by considering the two proteins as a network, with residues as nodes and contacts as edges. i‐Patch starts with two proteins, A and B, which are assumed to interact, but for which the structure of the complex is not available. However, we assume that for each protein, we have a reference structure and a multiple sequence alignment of homologues. i‐Patch then uses the propensities of patches of residues to interact, to predict interprotein contact sites. i‐Patch outperforms several other tested algorithms for prediction of interprotein contact sites. It gives 59% precision with 20% recall on a blind test set of 31 protein pairs. Combining the i‐Patch scores with an existing correlated mutation algorithm, McBASC, using a logistic model gave little improvement. Results from a case study, on bacterial chemotaxis protein complexes, demonstrate that our predictions can identify contact residues, as well as suggesting unknown interfaces in multiprotein complexes. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

Protein domain interfaces: characterization and comparison with oligomeric protein interfaces

The physical and chemical properties of domain-domain interactions have been analysed in two-domain proteins selected from the protein classification, CATH. The two-domain structures were divided into those derived from (i) monomeric proteins, or (ii) oligomeric or complexed proteins. The size, polarity, hydrogen bonding and packing of the intra-chain domain interface were calculated for both sets of two-domain structures. The results were compared with inter-chain interface parameters from permanent and non-obligate protein-protein complexes. In general, the intra-chain domain and inter-chain interfaces were remarkably similar. Many of the intra-chain interface properties are intermediate between those calculated for permanent and non-obligate inter-chain complexes. Residue interface propensities were also found to be very similar, with hydrophobic residues playing a major role, together with positively charged arginine residues. In addition, the residue composition of the domain interfaces were found to be more comparable with domain surfaces than domain cores. The implications of these results for domain swapping and protein folding are discussed.     

Interaction of the interleukin 8 protein with a sodium dodecyl sulfate micelle: A computer simulation study

Molecular simulations were carried out to study the sodium dodecyl sulfate (SDS) surfactant with the interleukin 8 (IL8) protein as a model to investigate the influence of amphiphilic molecules on proteins. Simulations for an SDS micelle with an IL8 protein show that both aggregates, which were initially separated, eventually approach each other to form a single complex. The results showed that the protein was attached to the SDS micelle by the charged positive amino acids whereas less contacts were observed for the negatively charged amino acids. Structural protein properties, such as amino acid contacts and pair correlation functions were conducted between the micelle and the protein groups and they showed greater interactions between the surfactant headgroups and the positively charged residues in the protein. Moreover, hydrogen bonds were also calculated between both structures and a greater number of bonds among the SDS headgroups and the charged positive amino acids in the protein was found.

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Graphical Abstract Attachment of the interleukin 8 protein with a sodium dodecyl sulfate (SDS) micelle is given by the charged positive amino acids as indicated by the interaction of those amino acids with the SDS headgroups.

     

Recognition of nucleic acid bases and base-pairs by hydrogen bonding to amino acid side-chains

Sequence-specific protein-nucleic acid recognition is determined, in part, by hydrogen bonding interactions between amino acid side-chains and nucleotide bases. To examine the repertoire of possible interactions, we have calculated geometrically plausible arrangements in which amino acids hydrogen bond to unpaired bases, such as those found in RNA bulges and loops, or to the 53 possible RNA base-pairs. We find 32 possible interactions that involve two or more hydrogen bonds to the six unpaired bases (including protonated A and C), 17 of which have been observed. We find 186 "spanning" interactions to base-pairs in which the amino acid hydrogen bonds to both bases, in principle allowing particular base-pairs to be selectively targeted, and nine of these have been observed. Four calculated interactions span the Watson-Crick pairs and 15 span the G:U wobble pair, including two interesting arrangements with three hydrogen bonds to the Arg guanidinum group that have not yet been observed. The inherent donor-acceptor arrangements of the bases support many possible interactions to Asn (or Gln) and Ser (or Thr or Tyr), few interactions to Asp (or Glu) even though several already have been observed, and interactions to U (or T) only if the base is in an unpaired context, as also observed in several cases. This study highlights how complementary arrangements of donors and acceptors can contribute to base-specific recognition of RNA, predicts interactions not yet observed, and provides tools to analyze proposed contacts or design novel interactions.