首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 468 毫秒
1.
Role of cation-pi interactions to the stability of thermophilic proteins   总被引:3,自引:0,他引:3  
Elucidating the factors responsible for exhibiting extreme thermal stability of thermophilic proteins is very important for an understanding of the mechanism of protein stability, as well as to design stable proteins. In this work, we have analyzed the influence of cation-pi interactions to enhance the stability from mesophilic to thermophilic proteins. The favorable residue pairs forming such a system of interactions have been brought out. We found that the Tyr has a greater number of such interactions with Lys in thermophilic proteins. Specifically, the same Lys would experience a greater number of cation-pi interactions with several Tyr residues in thermophiles. On the other hand, the influence of Phe in making cation-pi interactions is higher in mesophiles than in thermophiles. Further, a network of cation-pi interactions are maintained by Lys in thermophiles, whereas Arg plays a major role in mesophilic proteins. Moreover, atoms that have a substantial positive charge in both Lys and Arg make a more significant contribution for cation-pi interactions than do cationic group atoms.  相似文献   

2.
We have investigated the roles played by C-Hcdots, three dots, centeredpi interactions in RNA binding proteins. There was an average of 55 C-Hcdots, three dots, centeredpi interactions per protein and also there was an average of one significant C-Hcdots, three dots, centeredpi interaction for every nine residues in the 59 RNA binding proteins studied. Main-chain to side-chain C-Hcdots, three dots, centeredpi interactions is the predominant type of interactions in RNA binding proteins. The donor atom contribution to C-Hcdots, three dots, centeredpi interactions was mainly from Phe, Tyr, Trp, Pro, Gly, Lys, His and Ala residues. The acceptor atom contribution to main-chain to side-chain C-Hcdots, three dots, centeredpi and side-chain to side-chain C-Hcdots, three dots, centeredpi interactions was mainly from Phe and Tyr residues. On the contrary, the acceptor atoms of Trp residues contributed to all the four types of C-Hcdots, three dots, centeredpi interactions. Also, Trp contributed both donor and acceptor atoms in main-chain to side-chain, main-chain to side-chain five-member aromatic ring and side-chain to side-chain C-Hcdots, three dots, centeredpi interactions. The secondary structure preference analysis of C-Hcdots, three dots, centeredpi interacting residues showed that, Arg, Gln, Glu, His, Ile, Leu, Lys, Met, Phe and Tyr preferred to be in helix, while Ala, Asp, Cys, Gly, Trp and Val preferred to be in strand conformation. Long-range C-Hcdots, three dots, centeredpi interactions are the predominant type of interactions in RNA binding proteins. More than 50% of C-Hcdots, three dots, centeredpi interacting residues had a higher conservation score. Significant percentage of C-Hcdots, three dots, centeredpi interacting residues had one or more stabilization centers. Seven percent of the theoretically predicted stabilizing residues were also involved in C-Hcdots, three dots, centeredpi interactions and hence these residues may also contribute additional stability to RNA binding proteins.  相似文献   

3.
Factors enhancing protein thermostability   总被引:22,自引:0,他引:22  
  相似文献   

4.
Proteins from thermophilic organisms exhibit high thermal stability, but have structures that are very similar to their mesophilic homologues. In order to gain insight into the basis of thermostability, we have analyzed the medium- and long-range contacts in mesophilic and thermophilic proteins of 16 different families. We found that the thermophiles prefer to have contacts between residues with hydrogen-bond-forming capability. Apart from hydrophobic contacts, more contacts are observed between polar and non-polar residues in thermophiles than mesophiles. Residue-wise analysis showed that Tyr has good contacts with several other residues, and Cys has considerably higher long-range contacts in thermophiles compared with mesophiles. Furthermore, the residues occurring in the range of 31-34 residues apart in the sequence contribute significant long-range contacts to the stability of thermophilic proteins.  相似文献   

5.
We attempt to understand the origin of enhanced stability in thermophilic proteins by analyzing thermodynamic data for 116 proteins, the largest data set achieved to date. We compute changes in entropy and enthalpy at the convergence temperature where different driving forces are maximally decoupled, in contrast to the majority of previous studies that were performed at the melting temperature. We find, on average, that the gain in enthalpy upon folding is lower in thermophiles than in mesophiles, whereas the loss in entropy upon folding is higher in mesophiles than in thermophiles. This implies that entropic stabilization may be responsible for the high melting temperature, and hints at residual structure or compactness of the denatured state in thermophiles. We find a similar trend by analyzing a homologous set of proteins classified based only on the optimum growth temperature of the organisms from which they were extracted. We find that the folding free energy at the temperature of maximal stability is significantly more favorable in thermophiles than in mesophiles, whereas the maximal stability temperature itself is similar between these two classes. Furthermore, we extend the thermodynamic analysis to model the entire proteome. The results explain the high optimal growth temperature in thermophilic organisms and are in excellent quantitative agreement with full thermal growth rate data obtained in a dozen thermophilic and mesophilic organisms.  相似文献   

6.
We address the question of the thermal stability of proteins in thermophiles through comprehensive genome comparison, focussing on the occurrence of salt bridges. We compared a set of 12 genomes (from four thermophilic archaeons, one eukaryote, six mesophilic eubacteria, and one thermophilic eubacteria). Our results showed that thermophiles have a greater content of charged residues than mesophiles, both at the overall genomic level and in alpha helices. Furthermore, we found that in thermophiles the charged residues in helices tend to be preferentially arranged with a 1–4 helical spacing and oriented so that intra-helical charge pairs agree with the helix dipole. Collectively, these results imply that intra-helical salt bridges are more prevalent in thermophiles than mesophiles and thus suggest that they are an important factor stabilizing thermophilic proteins. We also found that the proteins in thermophiles appear to be somewhat shorter than those in mesophiles. However, this later observation may have more to do with evolutionary relationships than with physically stabilizing factors. In all our statistics we were careful to controls for various biases. These could have, for instance, arisen due to repetitive or duplicated sequences. In particular, we repeated our calculation using a variety of random and directed sampling schemes. One of these involved making a "stratified sample," a representative cross-section of the genomes derived from a set of 52 orthologous proteins present roughly once in each genome. For another sample, we focused on the subset of the 52 orthologs that had a known 3D structure. This allowed us to determine the frequency of tertiary as well as main-chain salt bridges. Our statistical controls supported our overall conclusion about the prevalence of salt bridges in thermophiles in comparison to mesophiles. Electronic Publication  相似文献   

7.
Lin YS 《Proteins》2008,73(1):53-62
Factors that are related to thermostability of proteins have been extensively studied in recent years, especially by comparing thermophiles and mesophiles. However, most of them are global characters. It is still not clear how to identify specific residues or fragments which may be more relevant to protein thermostability. Moreover, some of the differences among the thermophiles and mesophiles may be due to phylogenetic differences instead of thermal adaptation. To resolve these problems, I adopted a strategy to identify residue substitutions evolved convergently in thermophiles or mesophiles. These residues may therefore be responsible for thermal adaptation. Four classes of genomes were utilized in this study, including thermophilic archaea, mesophilic archaea, thermophilic bacteria, and mesophilic bacteria. For most clusters of orthologous groups (COGs) with sequences from all of these four classes of genomes, I can identify specific residues or fragments that may potentially be responsible for thermal adaptation. Functional or structural constraints (represented as sequence conservation) were suggested to have higher impact on thermal adaptation than secondary structure or solvent accessibility does. I further compared thermophilic archaea and mesophilic bacteria, and found that the most diverged fragments may not necessarily correspond to the thermostability-determining ones. The usual approach to compare thermophiles and mesophiles without considering phylogenetic relationships may roughly identify sequence features contributing to thermostability; however, to specifically identify residue substitutions responsible for thermal adaptation, one should take sequence evolution into consideration.  相似文献   

8.
The factors contributing to the thermal stability of proteins from thermophilic origins are matters of intense debate and investigation. Thermophilic proteins are thought to possess better packed interiors than their mesophilic counterparts, leading to lesser overall flexibility and a corresponding reduction in surface-to-volume ratio. These observations prompted an analysis of B values reported in high-resolution X-ray crystal structures of mesophilic and thermophilic proteins. In this analysis, the following aspects were addressed: (1) frequency distribution of normalized B values (B' factors) over all the proteins and for individual amino acids; (2) amino acid compositions in high B value regions of polypeptide chains; (3) variation in the B values from core to the surface of proteins in terms of their radius of gyration; and (4) degree of dispersion of normalized B values in spheres around the Calpha atoms. The analysis revealed that (1) Ser and Thr have lesser flexibility in thermophiles than in mesophiles, (2) the proportion of Glu and Lys in high B value regions of thermophiles is higher and that of Ser and Thr is lower and (3) the dispersion of B values within spheres at Calpha atoms is similar in mesophiles and thermophiles. These observations reflect plausible differences in the dynamics of thermophilic and mesophilic proteins and suggest amino acid substitutions that are likely to change thermal stability.  相似文献   

9.
Understanding the molecular basis for the enhanced stability of proteins from thermophiles has been hindered by a lack of structural data for homologous pairs of proteins from thermophiles and mesophiles. To overcome this difficulty, complete genome sequences from 9 thermophilic and 21 mesophilic bacterial genomes were aligned with protein sequences with known structures from the protein data bank. Sequences with high homology to proteins with known structures were chosen for further analysis. High quality models of these chosen sequences were obtained using homology modeling. The current study is based on a data set of models of 900 mesophilic and 300 thermophilic protein single chains and also includes 178 templates of known structure. Structural comparisons of models of homologous proteins allowed several factors responsible for enhanced thermostability to be identified. Several statistically significant, specific amino acid substitutions that occur going from mesophiles to thermophiles are identified. Most of these are at solvent-exposed sites. Salt bridges occur significantly more often in thermophiles. The additional salt bridges in thermophiles are almost exclusively in solvent-exposed regions, and 35% are in the same element of secondary structure. Helices in thermophiles are stabilized by intrahelical salt bridges and by an increase in negative charge at the N-terminus. There is an approximate decrease of 1% in the overall loop content and a corresponding increase in helical content in thermophiles. Previously overlooked cation-pi interactions, estimated to be twice as strong as ion-pairs, are significantly enriched in thermophiles. At buried sites, statistically significant hydrophobic amino acid substitutions are typically consistent with decreased side chain conformational entropy.  相似文献   

10.
Ribosomes were isolated from three mesophilic and three thermophilic strains of Bacillus. The ribosomes consisted of about 55% protein and 45% ribonucleic acid. Average ratios for the absorbance at 260/235 and 260/280 mmu were 1.77 and 1.92 for the mesophiles and 1.63 and 1.84 for the thermophiles. Ultracentrifugation revealed mainly components with sedimentation coefficients of about 30, 50, 70, 100, and 120S. All the preparations were shown to contain a ribonuclease which, in the presence of ethylenediaminetetraacetic acid, led to ribosome breakdown as measured by the increase in acid-soluble nucleotides. The stability of the ribosomes from the thermophiles was consistently greater than that of the ribosomes from the mesophiles. After 5 hr at 37 C, the breakdown was about 80% for the ribosomes from the mesophiles and 55 to 70% for those from the thermophiles. At 60 C, the ribosomes from the mesophiles were broken down slightly more and at a faster rate than those from the thermophiles. At temperatures above 60 C, the breakdown was again more pronounced for the ribosomes from the mesophiles.  相似文献   

11.
Liang HK  Huang CM  Ko MT  Hwang JK 《Proteins》2005,59(1):58-63
Structural analysis is useful in elucidating structural features responsible for enhanced thermal stability of proteins. However, due to the rapid increase of sequenced genomic data, there are far more protein sequences than the corresponding three-dimensional (3D) structures. The usual sequence-based amino acid composition analysis provides useful but simplified clues about the amino acid types related to thermal stability of proteins. In this work, we developed a statistical approach to identify the significant amino acid coupling sequence patterns in thermophilic proteins. The amino acid coupling sequence pattern is defined as any 2 types of amino acids separated by 1 or more amino acids. Using this approach, we construct the rho profiles for the coupling patterns. The rho value gives a measure of the relative occurrence of a coupling pattern in thermophiles compared with mesophiles. We found that thermophiles and mesophiles exhibit significant bias in their amino acid coupling patterns. We showed that such bias is mainly due to temperature adaptation instead of species or GC content variations. Though no single outstanding coupling pattern can adequately account for protein thermostability, we can use a group of amino acid coupling patterns having strong statistical significance (p values < 10(-7)) to distinguish between thermophilic and mesophilic proteins. We found a good correlation between the optimal growth temperatures of the genomes and the occurrences of the coupling patterns (the correlation coefficient is 0.89). Furthermore, we can separate the thermophilic proteins from their mesophilic orthologs using the amino acid coupling patterns. These results may be useful in the study of the enhanced stability of proteins from thermophiles-especially when structural information is scarce. Proteins 2005. (c) 2005 Wiley-Liss, Inc.  相似文献   

12.
Gromiha MM  Suresh MX 《Proteins》2008,70(4):1274-1279
Discriminating thermophilic proteins from their mesophilic counterparts is a challenging task and it would help to design stable proteins. In this work, we have systematically analyzed the amino acid compositions of 3075 mesophilic and 1609 thermophilic proteins belonging to 9 and 15 families, respectively. We found that the charged residues Lys, Arg, and Glu as well as the hydrophobic residues, Val and Ile have higher occurrence in thermophiles than mesophiles. Further, we have analyzed the performance of different methods, based on Bayes rules, logistic functions, neural networks, support vector machines, decision trees and so forth for discriminating mesophilic and thermophilic proteins. We found that most of the machine learning techniques discriminate these classes of proteins with similar accuracy. The neural network-based method could discriminate the thermophiles from mesophiles at the five-fold cross-validation accuracy of 89% in a dataset of 4684 proteins. Moreover, this method is tested with 325 mesophiles in Xylella fastidosa and 382 thermophiles in Aquifex aeolicus and it could successfully discriminate them with the accuracy of 91%. These accuracy levels are better than other methods in the literature and we suggest that this method could be effectively used to discriminate mesophilic and thermophilic proteins.  相似文献   

13.
The role of kinetics in governing exceptional stability of thermophilic ferredoxins has been explored. Strikingly, unfolding-rate constants (pH 7, 20 degrees C) are over eight (log(10)) orders of magnitude slower for the thermophiles than for a large set of unrelated mesophilic proteins. Also at low pH, where ionic interactions are diminished, unfolding of the thermophilic ferredoxins is significantly slower than unfolding of the mesophiles at pH 7, emphasizing the importance of hydrophobic interactions. A kinetic barrier towards unfolding may be a common strategy used by many proteins to withstand extreme conditions.  相似文献   

14.
Singer GA  Hickey DA 《Gene》2003,317(1-2):39-47
A number of recent studies have shown that thermophilic prokaryotes have distinguishable patterns of both synonymous codon usage and amino acid composition, indicating the action of natural selection related to thermophily. On the other hand, several other studies of whole genomes have illustrated that nucleotide bias can have dramatic effects on synonymous codon usage and also on the amino acid composition of the encoded proteins. This raises the possibility that the thermophile-specific patterns observed at both the codon and protein levels are merely reflections of a single underlying effect at the level of nucleotide composition. Moreover, such an effect at the nucleotide level might be due entirely to mutational bias. In this study, we have compared the genomes of thermophiles and mesophiles at three levels: nucleotide content, codon usage and amino acid composition. Our results indicate that the genomes of thermophiles are distinguishable from mesophiles at all three levels and that the codon and amino acid frequency differences cannot be explained simply by the patterns of nucleotide composition. At the nucleotide level, we see a consistent tendency for the frequency of adenine to increase at all codon positions within the thermophiles. Thermophiles are also distinguished by their pattern of synonymous codon usage for several amino acids, particularly arginine and isoleucine. At the protein level, the most dramatic effect is a two-fold decrease in the frequency of glutamine residues among thermophiles. These results indicate that adaptation to growth at high temperature requires a coordinated set of evolutionary changes affecting (i) mRNA thermostability, (ii) stability of codon-anticodon interactions and (iii) increased thermostability of the protein products. We conclude that elevated growth temperature imposes selective constraints at all three molecular levels: nucleotide content, codon usage and amino acid composition. In addition to these multiple selective effects, however, the genomes of both thermophiles and mesophiles are often subject to superimposed large changes in composition due to mutational bias.  相似文献   

15.
It has long been known that amino acid substitutions in proteins of organisms living at moderate and high temperatures (mesophiles and thermophiles, respectively) are not all symmetrical; for example, more aligned sites have lysine in mesophiles and arginine in thermophiles than have the opposite pattern. This is generally taken to indicate that certain amino acids are favored over others by selection at different temperatures. Previous comparisons of protein sequences from mesophiles and thermophiles have used relatively small numbers of sequences from a diverse array of species, meaning that only the most common amino acid substitutions could be examined and any taxon-specific patterns would be obscured. Here, we compare a large number of proteins between mesophiles and thermophiles in the archaeal genus Methanococcus and the bacterial genus Bacillus. Each genus exhibits dramatically asymmetrical substitution patterns for many pairs of amino acids. There are several pairs of amino acids for which one amino acid is favored in thermophilic Bacillus and the other is favored in thermophilic Methanococcus; this appears to result from the higher G + C content of the DNA of thermophilic Bacillus, a complication not seen in Methanococcus.  相似文献   

16.
Human recombinant granulocyte-colony stimulating factor (rhG-CSF) has one free cysteine at position 17 and has two disulfide bridges (Cys36-Cys42 and Cys64-Cys74). The Cys17 of rhG-CSF was substituted with Gly, Ala, Ser, Ile, Tyr, Arg, and Pro, or deleted using site-directed mutagenesis in order to improve its thermostability. With the exception of Pro17-rhG-CSF, all mutant proteins retained biological activity which promotes the growth of mouse bone marrow cells in vitro. Among these mutant proteins, Ala17-rhG-CSF had more than 5 times higher stability than rhG-CSF. But Ser17-rhG-CSF had almost same stability as rhG-CSF and other mutant proteins had only lower stability.  相似文献   

17.
The unparalleled peroxisome-to-mitochondrion mistargeting of alanine:glyoxylate aminotransferase (AGT) in the hereditary disease primary hyperoxaluria type 1 is caused by the combined presence of a common Pro11 --> Leu polymorphism and a disease-specific Gly170 --> Arg mutation. The Pro11 --> Leu replacement generates a functionally weak N-terminal mitochondrial targeting sequence (MTS), the efficiency of which is increased by the additional presence of the Gly170 --> Arg replacement. AGT dimerization is inhibited in the combined presence of both replacements but not when each is present separately. In this paper we have attempted to identify the structural determinants of AGT dimerization and mitochondrial mistargeting. Unlike most MTSs, the polymorphic MTS of AGT has little tendency to adopt an alpha-helical conformation in vitro. Nevertheless, it is able to target efficiently a monomeric green fluorescent (GFP) fusion protein, but not dimeric AGT, to mitochondria in transfected COS-1 cells. Increasing the propensity of this MTS to fold into an alpha-helix, by making a double Pro11 --> Leu + Pro10 --> Leu replacement, enabled it to target both GFP and AGT efficiently to mitochondria. The double Pro11 --> Leu + Pro10 --> Leu replacement retarded AGT dimerization in vitro as did the disease-causing double Pro11 --> Leu + Gly170 --> Arg replacement. These data suggest that N-terminal alpha-helix formation is more important for maintaining AGT in a conformation (i. e. monomeric) compatible with mitochondrial import than it is for the provision of mitochondrial targeting information. The parallel effects of the Pro10 --> Leu and Gly170 --> Arg replacements on the dimerization and intracellular targeting of polymorphic AGT (containing the Pro11 --> Leu replacement) raise the possibility that they might achieve their effects by the same mechanism.  相似文献   

18.
Abstract: The molecular origin of protein stability has been the subject of active research for more than a generation (R. Jaenicke (1991) Eur. J. Biochem. 202, 715–728). Faced with the discovery of extremophiles, in recent years the problem has gained momentum, especially because of its biotechnological potential. In analyzing a number of enzymes from the hyperthermophilic bacterium Thermotoga maritima , it has become clear that the excess free energy of stabilization is equivalent to only a few weak bonds ( ΔΔG stab≈ 50 kJ/mol). As taken from the comparison of homologous enzymes from mesophiles, thermophiles and hyperthermophiles, these accumulate from local interactions (especially ion pairs), enhanced secondary or supersecondary structure, and improved packing of domains and/or subunits, without significantly altering the overall topology. In this review, glyceraldehyde-3-phosphate dehydrogenase will be discussed as a representative example to illustrate possible adaptive strategies to the extreme thermal stress in hydrothermal vents.  相似文献   

19.
Proteins are multifunctional: their amino acid sequences simultaneously determine folding, function and turnover. Correspondingly, evolution selected for compromises between rigidity (stability) and flexibility (folding/function/degradation), to the result that generally the free energy of stabilization of globular proteins in solution is the equivalent to only a few weak intermolecular interactions. Additional increments may come from extrinsic factors such as ligands or specific compatible solutes. Apart from the enthalpic effects, entropy may play a role by reducing the flexibility (cystine bridges, increased proline content), or by water release from residues buried upon folding and association. Additional quaternary interactions and closer packing are typical characteristics of proteins from thermophiles. In halophiles, protein stability and function are maintained by increased ion binding and glutamic acid content, both allowing the protein inventory to compete for water at high salt. Acidophiles and alkalophiles show neutral intracellular pH; proteins facing the outside extremes of pH possess anomalously high contents in ionizable amino acids. Global comparisons of the amino acid compositions and sequences of proteins from mesophiles and extremophiles did not result in general rules of protein stabilization, even after including complete genome sequences into the search. Obviously, proteins are individuals that optimize internal packing and external solvent interactions by very different mechanisms, each protein in its own way. Strategies deduced from specific ultrastable proteins allow stabilizing point mutations to be predicted.  相似文献   

20.
Seven hyper-stable multiple mutants have been constructed in staphylococcal nuclease by various combinations of eight different stabilizing single mutants. The stabilities of these multiple mutants determined by guanidine hydrochloride denaturation were 3.4 to 5.6 kcal/mol higher than that of the wild-type. Their thermal denaturation midpoint temperatures were 12.6 to 22.9 deg. C higher than that of the wild-type. These are among the greatest increases in protein stability and thermal denaturation midpoint temperature relative to the wild-type yet attained. There has been great interest in understanding how proteins found in thermophilic organisms are stabilized. One frequently cited theory is that the packing of hydrophobic side-chains is improved in the cores of proteins isolated from thermophiles when compared to proteins from mesophiles. The crystal structures of four single and five multiple stabilizing mutants of staphylococcal nuclease were solved to high resolution. No large overall structural change was found, with most changes localized around the sites of mutation. Rearrangements were observed in the packing of side-chains in the major hydrophobic core, although none of the mutations was in the core. It is surprising that detailed structural analysis showed that packing had improved, with the volume of the mutant protein's hydrophobic cores decreasing as protein stability increased. Further, the number of van der Waals interactions in the entire protein showed an experimentally significant increase correlated with increasing stability. These results indicate that optimization of packing follows as a natural consequence of increased protein thermostability and that good packing is not necessarily the proximate cause of high stability. Another popular theory is that thermostable proteins have more electrostatic and hydrogen bonding interactions and these are responsible for the high stabilities. The mutants here show that increased numbers of electrostatic and hydrogen bonding interactions are not obligatory for large increases in protein stability.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号