Determinants of strand register in antiparallel beta-sheets of proteins.

Antiparallel beta-sheets present two distinct environments to inter-strand residue pairs: beta(A,HB) sites have two backbone hydrogen bonds; whereas at beta(A,NHB) positions backbone hydrogen bonding is precluded. We used statistical methods to compare the frequencies of amino acid pairs at each site. Only approximately 10% of the 210 possible pairs showed occupancies that differed significantly between the two sites. Trends were clear in the preferred pairs, and these could be explained using stereochemical arguments. Cys-Cys, Aromatic-Pro, Thr-Thr, and Val-Val pairs all preferred the beta(A,NHB) site. In each case, the residues usually adopted sterically favored chi1 conformations, which facilitated intra-pair interactions: Cys-Cys pairs formed disulfide bonds; Thr-Thr pairs made hydrogen bonds; Aromatic-Pro and Val-Val pairs formed close van der Waals contacts. In contrast, to make intimate interactions at a beta(A,HB) site, one or both residues had to adopt less favored chi1 geometries. Nonetheless, pairs containing glycine and/or aromatic residues were favored at this site. Where glycine and aromatic side chains combined, the aromatic residue usually adopted the gauche conformation, which promoted novel aromatic ring-peptide interactions. This work provides rules that link protein sequence and tertiary structure, which will be useful in protein modeling, redesign, and de novo design. Our findings are discussed in light of previous analyses and experimental studies.     

Statistical analysis of predominantly transient protein-protein interfaces

A non-redundant set of 170 protein-protein interfaces of known structure was statistically analyzed for residue and secondary-structure compositions, pairing preferences and side-chain-backbone interaction frequencies. By focussing mainly on transient protein-protein interfaces, the results underline previous findings for protein-protein interfaces but also show some new interesting aspects of transient interfaces. The residue compositions at interfaces found in this study correlate well with the results of other studies. On average, contacts between pairs of hydrophobic and polar residues were unfavorable, and the charged residues tended to pair subject to charge complementarity. Secondary structure composition analysis shows that neither helices nor beta-sheets are dominantly populated at interfaces. Analyzing the pairing preferences of the secondary structure elements revealed a higher affinity within the same elements and alludes to tight packings. In addition, the results for the side-chain and backbone interaction frequencies, which were measured under more stringent conditions, showed a high occurrence of side-chain-backbone interactions. Taking a closer look at the helix and beta-sheet binding frequencies for a given side-chain and backbone interaction underlined the relevance of tight packings. The polarity of interfaces increased with decreasing interface size. These types of information may be useful for scoring complexes in protein-protein docking studies or for prediction of protein-protein interfaces from the sequences alone.     

The interrelationships of side-chain and main-chain conformations in proteins

The accurate determination of a large number of protein structures by X-ray crystallography makes it possible to conduct a reliable statistical analysis of the distribution of the main-chain and side-chain conformational angles, how these are dependent on residue type, adjacent residue in the sequence, secondary structure, residue-residue interactions and location at the polypeptide chain termini. The interrelationship between the main-chain (phi, psi) and side-chain (chi 1) torsion angles leads to a classification of amino acid residues that simplify the folding alphabet considerably and can be a guide to the design of new proteins or mutational studies. Analyses of residues occurring with disallowed main-chain conformation or with multiple conformations shed some light on why some residues are less favoured in thermophiles.     

Relationship between ion pair geometries and electrostatic strengths in proteins

The electrostatic free energy contribution of an ion pair in a protein depends on two factors, geometrical orientation of the side-chain charged groups with respect to each other and the structural context of the ion pair in the protein. Conformers in NMR ensembles enable studies of the relationship between geometry and electrostatic strengths of ion pairs, because the protein structural contexts are highly similar across different conformers. We have studied this relationship using a dataset of 22 unique ion pairs in 14 NMR conformer ensembles for 11 nonhomologous proteins. In different NMR conformers, the ion pairs are classified as salt bridges, nitrogen-oxygen (N-O) bridges and longer-range ion pairs on the basis of geometrical criteria. In salt bridges, centroids of the side-chain charged groups and at least a pair of side-chain nitrogen and oxygen atoms of the ion-pairing residues are within a 4 A distance. In N-O bridges, at least a pair of the side-chain nitrogen and oxygen atoms of the ion-pairing residues are within 4 A distance, but the distance between the side-chain charged group centroids is greater than 4 A. In the longer-range ion pairs, the side-chain charged group centroids as well as the side-chain nitrogen and oxygen atoms are more than 4 A apart. Continuum electrostatic calculations indicate that most of the ion pairs have stabilizing electrostatic contributions when their side-chain charged group centroids are within 5 A distance. Hence, most (approximately 92%) of the salt bridges and a majority (68%) of the N-O bridges are stabilizing. Most (approximately 89%) of the destabilizing ion pairs are the longer-range ion pairs. In the NMR conformer ensembles, the electrostatic interaction between side-chain charged groups of the ion-pairing residues is the strongest for salt bridges, considerably weaker for N-O bridges, and the weakest for longer-range ion pairs. These results suggest empirical rules for stabilizing electrostatic interactions in proteins.     

Specific sequence of motifs of mitochondrial uncoupling proteins

We have searched for the exclusivity of common sequence motifs of the mitochondrial uncoupling proteins (UCP1, UCP2, UCP3, UCP4, BMCP1, and plant UCP [PUMP]) within the gene family of mitochondrial anion carrier proteins. The UCP-specific sequences, "UCP signatures", were found in the first, second, and fourth alpha-helices. First: Ala/Ser-Cys/Thr/n-n/Phe-Ala/Gly-[negatively charged residue]-n/Phe-n/Cys-Thr-Phe/n; second: Gly/Ala-Ile/Leu-Gln/X-[positively charged residue]-NH-n/Cys-Ser/nphi/X-n/Ser-OH/Gly-n-[positively charged residue]-Ile/Met-Gly/Val-n/Thr; fourth: Pro-Asn/ Thr-n-X-[positively charged residue]-Asn/Ser/Ala-n-n-Ile/Leu-n-Asn/Val-Cys/n-n/Thr-[negatively charged residue]-n-n/Thr/Pro-OH/Val (n, nonpolar; phi, aromatic; (positively charged residue/negatively charged residue, charged residue). The second and part of the third signature are also present in the yeast dicarboxylate transporter. The UCP signature excluding BMCP1 was also found in the second matrix segment: [positively charged residue]-(Pro/ del-Leu/del)-[positively charged residue]-phi-X-Gly/Ser-Thr/n-X-NH/[negatively charged residue]-Ala-phi. These UCP signatures are thought to be involved in fatty acid anion binding and translocation.     

Thermo- and Mesostabilizing Protein Interactions Identified by Temperature-Dependent Statistical Potentials

The goal of controlling protein thermostability is tackled here through establishing, by in silico analyses, the relative weight of residue-residue interactions in proteins as a function of temperature. We have designed for that purpose a (melting-) temperature-dependent, statistical distance potential, where the interresidue distances are computed between the side-chain geometric centers or their functional centers. Their separate derivation from proteins of either high or low thermal resistance reveals the interactions that contribute most to stability in different temperature ranges. Thermostabilizing interactions include salt bridges and cation-π interactions (especially those involving arginine), aromatic interactions, and H-bonds between negatively charged and some aromatic residues. In contrast, H-bonds between two polar noncharged residues or between a polar noncharged residue and a negatively charged residue are relatively less stabilizing at high temperatures. An important observation is that it is necessary to consider both repulsive and attractive interactions in overall thermostabilization, as the degree of repulsion may also vary with temperature. These temperature-dependent potentials are not only useful for the identification of meso- and thermostabilizing pair interactions, but also exhibit predictive power, as illustrated by their ability to predict the melting temperature of a protein based on the melting temperature of homologous proteins.     

Energetics of charge-charge interactions between residues adjacent in sequence

Statistical analysis of the residue separation between a pair of ionizable side chains within 4 ? of each other was performed on a set of 1560 non-homologous PDB structures. We found that the frequency of pairs of like charges (i.e., pairs consisting of acidic residues Asp and Glu or pairs consisting of basic residues Arg and Lys) is two orders of magnitude lower than the pairs of oppositely charged residues (salt-bridges). We also found that for pairs of like charges the distribution is skewed dramatically towards short residue separation (<3). On the basis of these observations, we hypothesize that at short residue separation the repulsion between charges does not contribute much to the protein stability and the effects are largely dominated by the long range charge-charge interactions with other ionizable groups in the protein molecule. To test this hypothesis, we incorporated various pairs of charged residues at position 63 and 64 of ubiquitin and compared the stabilities of these variants. We also performed calculations of the expected changes in the charge-charge interactions. A very good correlation between experimental changes in the stability of ubiquitin variants, and changes in the energy of charge-charge interactions provides support for the hypothesis that a pair of ionizable residues next to each other in sequence modulates protein stability via long range charge-charge interactions with the rest of the protein.     

Analysis of side-chain orientations in homologous proteins

The side-chain conformations of topologically equivalent residues in seven pairs of proteins ranging in sequence homology from 16% to 60% are compared. Both identical and mutated residues are included. For proteins with greater than 40% homology, it is found that at least 80% of the side-chain orientations of identical residues and 75% or more of the mutated residues in each pair of proteins have matching gamma atom dihedral angles (+/- 40 degrees); the comparison is not based strictly on chi 1 angles. Further, if a match is obtained at the gamma position, there is a high probability of matching for the delta atom(s) of the side-chain. For proteins with less than 25% homology the percentages are somewhat lower. Trends observed for conservative substitutions are essentially the same as those noted for mutated residues in general. Side-chain accessibility does not affect the probability of matches of identical residues; however, less accessible pairs of mutated residues have 10 to 20% higher matching probabilities than do exposed residues. Mismatches can frequently be related to large B-factors, certain types of amino acid substitutions, or the appearance of multiple minima on the side-chain potential energy surfaces and are most likely to occur for certain small residues (Ser, Thr, Val). Analysis of all the results makes possible the formulation of a set of rules for side-chain positioning in the modeling of homologous proteins.     

Comparative study of the phenylalanyl-tRNA synthetases from Escherichia coli and Thermus thermophlus by the tritium topography method

A comparative study of thermostability and aminoacid composition of the phenylalanyl-tRNA synthetases from E. coli and Thermus thermophilus HB8 has been carried out. Compared with the mesophilic enzyme, a considerable increase of Pro, Leu, Phe, Arg and decrease of Asx, Ile, Ser, Thr and Lys content have been revealed in the thermophilic protein. Using tritium topography, Pro, (Leu + Ile) and Gly were found to be the most accessible on the surfaces of both the enzymes. In the E. coli enzyme, Thr residues were also easy to access while on the surface of the thermophilic enzyme there were more Arg residues. The quantitative assay of the surface compositions revealed the increased exposure of the (Leu + Ile) residues on the thermophilic protein as well as of the charged Asx and Arg residues. A possible correlation of the observed effects with thermostability is discussed.     

On residues in the disallowed region of the Ramachandran map.

An analysis of the occurrence of nonglycyl residues in conformations disallowed in the Ramachandran plot is presented. Ser, Asn, Thr, and Cys have the highest propensities to exhibit such conformations, and the branched aliphatic residues the lowest. Residues cluster in five regions and there are some trends in the types of residues and their side-chain conformations (chi(1)) occupying these. Majority of the residues are found at the edge of helices and strands and in short loops, and are involved in different types of weak, stabilizing interactions. A structural motif has been identified where a residue in disallowed conformation occurs as the first residue of a short 3(10)-helix. On the basis of the types of neighboring residues, the location in the three-dimensional structure and accessibility, there are similarities with the occurrence of cis peptide bonds in protein structures.     

Rearrangement of charge-charge interactions in variant ubiquitins as detected by double-mutant cycles and NMR

Previous studies of ubiquitin disclosed numerous charge-charge interactions on the protein's surface. To investigate how neighboring residues influence the strength of these interactions, double-mutant cycles are combined with pK(a) determinations by 2D NMR. More specifically, the environment around the Asp21-Lys29 ion pair has been altered through mutations at position 25, which is an asparagine in mammalian ubiquitin and a positively-charged residue in many other ubiquitin-like proteins. The pK(a) value of Asp21 decreases by 0.4 to 0.7 pH unit when Asn25 is substituted with a positively charged residue, suggesting a new and favorable ion pair interaction between positions 21 and 25. However, analysis of double mutants reveals that the favorable interaction between Asp21 and Lys29 is weakened when position 25 is a positively charged residue. Interestingly, while the pK(a) value of His25 in the N25H variant agrees with model compound values, additional mutants reveal that this agreement is fortuitous, resulting from a balance of favorable and unfavorable interactions; similar results were observed previously for Glu34 in ubiquitin and His8 in staphylococcal nuclease. Ionizable groups may thus have pK(a) values similar to model compound values and yet still be involved in significant interactions with other protein groups. One surprising result of introducing positively charged residues at position 25 is a new interaction between Lys29 and Glu18, an interaction not present in wild-type ubiquitin. This unanticipated result illustrates a key advantage of using NMR to determine pK(a) values for many residues simultaneously in the variant proteins. Overall, the strength of an interaction between two residues at the surface of ubiquitin is sensitive to the identity of neighboring residues. The results also demonstrate that relatively conservative and common point mutations such as substitutions of polar with charged residues and vice versa can have effects on interactions beyond the site of mutation per se.     

An analysis of side chain interactions and pair correlations within antiparallel β-sheets: The differences between backbone hydrogen-bonded and non-hydrogen-bonded residue pairs

Cross-strand pair correlations are calculated for residue pairs in antiparallel β-sheet for two cases: pairs whose backbone atoms are hydrogen bonded together (H-bonded site) and pairs which are not (non-H-bonded site). The statistics show that this distinction is important. When glycine is located on the edge of a sheet, it shows a 3:1 preference for the H-bonded site. Thestrongest observed correlations are for pairs of disulfide-bonded cystines, many of which adopt a close-packed conformation with each cystine in a spiral conformation of opposite chirality to its partner. It is likely that these pairs are a signature for the family of small, cystine-rich proteins. Most other strong positive and negative correlations involve charged and polar residues. It appears that electrostatic compatibility is the strongest factor affecting pair correlation. Significant correlations are observed for β- and γ-branched residues inthe non-H-bonded site. An examination of the structures showsa directionality in side chain packing. There is a correlation between (1) the directionality in the packing interactions of non-H-bonded β- and γ-branched residue pairs, (2) the handedness of the observed enantiomers of chiral β-branched side chains, and (3) the handedness of the twist of β-sheet. These findings have implications for the formation of β-sheets during protein folding and the mechanism by which the sheet becomes twisted. © 1995 Wiley-Liss, Inc.     

Anabaena flavodoxin as an electron carrier from photosystem I to ferredoxin-NADP+ reductase. Role of flavodoxin residues in protein-protein interaction and electron transfer

Biochemical and structural studies indicate that electrostatic and hydrophobic interactions are critical in the formation of optimal complexes for efficient electron transfer (ET) between ferredoxin-NADP(+) reductase (FNR) and ferredoxin (Fd). Moreover, it has been shown that several charged and hydrophobic residues on the FNR surface are also critical for the interaction with flavodoxin (Fld), although, so far, no key residue on the Fld surface has been found to be the counterpart of such FNR side chains. In this study, negatively charged side chains on the Fld surface have been individually modified, either by the introduction of positive charges or by their neutralization. Our results indicate that although Glu16, Glu20, Glu61, Asp65, and Asp96 contribute to the orientation and optimization of the Fld interaction, either with FNR or with photosystem I (PSI) (presumably through the formation of salt bridges), for efficient ET, none of these side chains is involved in the formation of crucial salt bridges for optimal interaction with FNR. These data support the idea that the FNR-Fld interaction is less specific than the FNR-Fd interaction. However, analysis of the reactivity of these mutated Flds toward the membrane-anchored PSI complex indicated that all mutants, except Glu16Gln, lack the ability to form a stable complex with PSI. Thr12, Thr56, Asn58, and Asn97 are present in the close environment of the isoalloxazine ring of FMN in Anabaena Fld. Their roles in the interaction with and ET to FNR and PSI have also been studied. Mutants at these Fld positions indicate that residues in the close environment of the isoalloxazine ring modulate the ability of Fld to bind to and to exchange electrons with its physiological counterparts.     

Role of neighboring FMN side chains in the modulation of flavin reduction potentials and in the energetics of the FMN:apoprotein interaction in Anabaena flavodoxin

Flavodoxins (Flds) are electron transfer proteins that carry a noncovalently bound flavin mononucleotide molecule (FMN) as a redox active center. A distinguishing feature of these flavoproteins is the dramatic change in the E(sq/rd) reduction potential of the FMN upon binding to the apoprotein (at pH 8.0, from -269 mV when free in solution to -438 mV in Anabaena Fld). In this study, the contribution of three neighboring FMN residues, Thr56, Asn58, and Asn97, and of three negatively charged surface residues, Glu20, Asp65, and Asp96, to modulate the redox properties of FMN upon its binding to the apoprotein has been investigated. Additionally, the role of these residues in the apoflavodoxin:FMN interaction has been analyzed. Concerning the redox potentials, the most noticeable result was obtained for the Thr56Gly mutant. In this Fld variant, the increased accessibility of FMN leads to an increase of +63 mV in the E(sq/rd) value. On the other hand, a correlation between the electrostatic environment of FMN and the E(sq/rd) has been observed. The more positive residues or the less negative residues present in the surroundings of the FMN N(1) atom, then the less negative the value for E(sq/rd). With regard to FMN binding to apoflavodoxin, breaking of hydrophobic interactions between FMN and residues 56, 58, and 97 seems to increase the K(d) values, especially in the Thr56Gly Fld. Such results suggest that the H-bond network in the FMN environment influences the FMN affinity.     

Beta2-microglobulin amyloid fragment organization and morphology and its comparison to Abeta suggests that amyloid aggregation pathways are sequence specific

Beta2-microglobulin (beta2-m) can form dialysis-related amyloid deposits. The structure of a fragment of beta2-m (K3, Ser20-Lys41) in the oligomeric state has recently been solved. We modeled equilibrium structures of K3 oligomers with different organizations (single and double layers) and morphologies (linear-like and annular-like) for the wild-type and mutants using all-atom molecular dynamics (MD) simulations. We focused on the sheet-to-sheet association force, which is the key in the amyloid organization and morphology. For the linear-like morphology, we observed two stable organizations: (i) single-layered parallel-stranded beta-sheets and (ii) double-layered parallel-stranded antiparallel beta-sheets stacked perpendicular to the fibril axis through the hydrophobic N-terminal-N-terminal (NN) interface. No stable annular structures were observed. The structural instability of the annular morphology was mainly attributed to electrostatic repulsion of three negatively charged residues (Asp15, Glu17, and Asp19) projecting from the same beta-strand surface. Linear-like and annular-like double-layered oligomers with the NN interface are energetically more favorable than other oligomers with C-terminal-C-terminal (CC) or C-terminal-N-terminal (CN) interfaces, emphasizing the importance of hydrophobic interactions and side-chain packing in stabilizing these oligomers. Moreover, only linear-like structures, rather than annular structures, with parallel beta-strands and antiparallel beta-sheet arrangements are possible intermediate states for the K3 beta2-m amyloid fibrils in solution. Comparing the beta2-m fragment with Abeta indicates that while both adopt similar beta-strand-turn-beta-strand motifs, the final amyloid structures can be dramatically different in size, structure, and morphology due to differences in side-chain packing arrangements, intermolecular driving forces, sequence composition, and residue positions, suggesting that the mechanism leading to distinct morphologies and the aggregation pathways is sequence specific.     

Fluctuations between stabilizing and destabilizing electrostatic contributions of ion pairs in conformers of the c-Myc-Max leucine zipper

In solution proteins often exhibit backbone and side-chain flexibility. Yet electrostatic interactions in proteins are sensitive to motions. Hence, here we study the contribution of ion pairs toward protein stability in a range of conformers which sample the conformational space in solution. Specifically, we focus on the electrostatic contributions of ion pairs to the stability of each of the conformers in the NMR ensemble of the c-Myc-Max leucine zipper and to their average energy minimized structure. We compute the electrostatic contributions of inter- and intra-helical ion pairs and of an ion pair network. We find that the electrostatic contributions vary considerably among the 40 NMR conformers. Each ion pair, and the network, fluctuates between being stabilizing and being destabilizing. This fluctation reflects the variability in the location of the ion pairing residues and in the geometric orientation of these residues, both with respect to each other and with respect to other charged groups in the rest of the protein. Ion pair interactions in the c-Myc-Max leucine zipper in solution depend on the protein conformer which is analyzed. Hence, the overall stabilizing (or destabilizing) contribution of an ion pair is conformer population-dependent. This study indicates that free energy calculations performed using the continuum electrostatics methodology are sensitive to protein conformational details.     

The G protein-coupled 5-HT1A receptor causes suppression of caspase-3 through MAPK and protein kinase Calpha

Several studies have analysed aromatic interactions, involving mostly phenylalanine, tyrosine and tryptophan. Only a few studies have considered histidine as an interacting aromatic residue. An extensive analysis of aromatic His-X interactions is performed here on a data set of 593 PDB structures: 68% of the histidine are involved in aromatic pairs and 1271 non-redundant His-X pairs were analysed. Thirty percent of these pairs involve an aromatic partner less than 6 apart in the sequence. These near-sequence pairs correspond to conformations which stabilise secondary structures, mainly alpha-helices when the residues are 4 apart and beta-strands when they are 2 apart in the sequence. The partners of the other His-X pairs (887, 70%) are more than 5 apart in the sequence. Of these far-sequence pairs, 35% bridge beta strands and only 9% helices. The near-sequence pairs are sterically constrained as supported by conformer distribution. The X partners of far-sequence His-X pairs are mainly "above" the histidine ring with tilted and normal rings, corresponding to a "T shape; face to edge" orientation. Phenylalanine, the only aromatic residue with no heteroatom, is a disfavoured partner, whereas histidine is the preferred one. Heteroatom-heteroatom interactions are favoured in near-sequence as well as in far-sequence His-His, His-Trp and His-Tyr pairs.     

Three-dimensional structures of thermophilic beta-1,4-xylanases from Chaetomium thermophilum and Nonomuraea flexuosa. Comparison of twelve xylanases in relation to their thermal stability.

The crystal structures of thermophilic xylanases from Chaetomium thermophilum and Nonomuraea flexuosa were determined at 1.75 and 2.1 A resolution, respectively. Both enzymes have the overall fold typical to family 11 xylanases with two highly twisted beta-sheets forming a large cleft. The comparison of 12 crystal structures of family 11 xylanases from both mesophilic and thermophilic organisms showed that the structures of different xylanases are very similar. The sequence identity differences correlated well with the structural differences. Several minor modifications appeared to be responsible for the increased thermal stability of family 11 xylanases: (a) higher Thr : Ser ratio (b) increased number of charged residues, especially Arg, resulting in enhanced polar interactions, and (c) improved stabilization of secondary structures involved the higher number of residues in the beta-strands and stabilization of the alpha-helix region. Some members of family 11 xylanases have a unique strategy to improve their stability, such as a higher number of ion pairs or aromatic residues on protein surface, a more compact structure, a tighter packing, and insertions at some regions resulting in enhanced interactions.     

Aromatic side-chain interactions in proteins. Near- and far-sequence His–X pairs

Several studies have analysed aromatic interactions, involving mostly phenylalanine, tyrosine and tryptophan. Only a few studies have considered histidine as an interacting aromatic residue. An extensive analysis of aromatic His–X interactions is performed here on a data set of 593 PDB structures: 68% of the histidine are involved in aromatic pairs and 1271 non-redundant His–X pairs were analysed. Thirty percent of these pairs involve an aromatic partner less than 6 apart in the sequence. These near-sequence pairs correspond to conformations which stabilise secondary structures, mainly α-helices when the residues are 4 apart and β-strands when they are 2 apart in the sequence. The partners of the other His–X pairs (887, 70%) are more than 5 apart in the sequence. Of these far-sequence pairs, 35% bridge beta strands and only 9% helices. The near-sequence pairs are sterically constrained as supported by conformer distribution. The X partners of far-sequence His–X pairs are mainly “above” the histidine ring with tilted and normal rings, corresponding to a “T shape; face to edge” orientation. Phenylalanine, the only aromatic residue with no heteroatom, is a disfavoured partner, whereas histidine is the preferred one. Heteroatom–heteroatom interactions are favoured in near-sequence as well as in far-sequence His–His, His–Trp and His–Tyr pairs.     

Beta-sheet capping: signals that initiate and terminate beta-sheet formation

In the present work, we address the question of whether different amino acids have different beta-sheet initiating and terminating characteristics. Using a large scale analysis of parallel and antiparallel beta-sheets in a non-redundant dataset of proteins, we observed that most of the amino acids show significant under- or over-representation in at least one of the positions at the two ends of beta-sheets, which are denoted as N-cap and C-cap. In addition, based on statistical data and structural comparison, we found that certain amino acids, especially Asp, Asn, Gly and Pro have strong tendencies to block beta-sheet continuation. Hence, we can consider these residues as beta-sheet terminators. It was also proposed that the dipole moments in parallel beta-sheets, whose direction is from C-terminal (partially negative) to N-terminal (partially positive), are much stronger than has previously been suggested. In fact, enhancement of dipole moments in parallel beta-sheets is a result of the positioning of positively charged residues at N-cap and negatively charged residues at C-cap. This enhancement in dipole moment magnitude leads to strengthened dipolar interactions between parallel beta-sheets dipoles and other partners especially alpha-helices dipoles. The results provide an explanation for the antiparallel alignment of parallel beta-sheets with alpha-helices.