Protein engineering最新文献_第10页

Common Structural Cliques: a tool for protein structure and function analysis. 共同结构团:蛋白质结构和功能分析的工具。

Protein engineering Pub Date : 2003-08-01 DOI: 10.1093/protein/gzg080

Mariusz Milik, Sándor Szalma, Krzysztof A Olszewski

{"title":"Common Structural Cliques: a tool for protein structure and function analysis.","authors":"Mariusz Milik, Sándor Szalma, Krzysztof A Olszewski","doi":"10.1093/protein/gzg080","DOIUrl":"https://doi.org/10.1093/protein/gzg080","url":null,"abstract":"Proposed is a method for locating functionally relevant atoms in protein structures and a representation of spatial arrangements of these atoms allowing for a flexible description of active sites in proteins. The search method is based on comparison of local structure features of proteins that share a common biochemical function. The method does not depend on overall similarity of structures and sequences of compared proteins or on previous knowledge about functionally relevant residues. The compared protein structures are condensed to a graph representation, with atoms as nodes and distances as edge labels. Protein graphs are then compared to extract all possible Common Structural Cliques. These cliques are merged to create Structural Templates: graphs that describe structural analogies between compared proteins. Structures of serine endopeptidases were compared in pairs using the presented algorithm with different geometrical parameters. Additionally, a Structural Template was extracted from the structures of aminotransferases, two different proteins that catalyze the same type of chemical reaction. The results presented show that the method works efficiently even in the case of large protein systems and allows for extraction of common structural features from proteins catalyzing a particular chemical reaction, but that evolved from different ancestors by convergent evolution.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 8","pages":"543-52"},"PeriodicalIF":0.0,"publicationDate":"2003-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg080","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22570766","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 52

Protein secondary structure prediction based on an improved support vector machines approach. 基于改进支持向量机方法的蛋白质二级结构预测。

Protein engineering Pub Date : 2003-08-01 DOI: 10.1093/protein/gzg072

Hyunsoo Kim, Haesun Park

引用次数: 228

Tandem repeats of Sushi3 peptide with enhanced LPS-binding and -neutralizing activities. 具有增强lps结合和中和活性的Sushi3肽串联重复序列。

Protein engineering Pub Date : 2003-08-01 DOI: 10.1093/protein/gzg078

Changgui Li, Miang Lon Patricia Ng, Yong Zhu, Bow Ho, Jeak Ling Ding

{"title":"Tandem repeats of Sushi3 peptide with enhanced LPS-binding and -neutralizing activities.","authors":"Changgui Li, Miang Lon Patricia Ng, Yong Zhu, Bow Ho, Jeak Ling Ding","doi":"10.1093/protein/gzg078","DOIUrl":"https://doi.org/10.1093/protein/gzg078","url":null,"abstract":"Endotoxin, also known as lipopolysaccharide (LPS), is the major mediator of septic shock due to Gram-negative bacterial infection. Chemically synthesized S3 peptide, derived from Sushi3 domain of Factor C, which is the endotoxin-sensitive serine protease of the limulus coagulation cascade, was previously shown to bind and neutralize LPS activity. For large-scale production of this peptide and to mimick other pathogen-recognizing molecules, tandem multimers of the S3 gene were constructed and expressed in Escherichia coli. The recombinant tetramer of S3 (rS3-4mer) was purified by anion exchange and digested into monomers (rS3-1mer). Both the rS3-4mer and rS3-1mer were functionally analyzed for their ability to bind LPS by an ELISA-based method and surface plasmon resonance. The LAL inhibition and TNFalpha-release test showed that rS3-1 mer can neutralize the LPS activity as effectively as the synthetic S3 peptide, while rS3-4mer displays an enhanced inhibitory effect on LPS-induced activities. Both recombinant peptides exhibited low cytotoxicity and no haemolytic activity on human cells. This evidence suggests that the recombinant sushi peptides have potential use for the detection, removal of endotoxin and/or anti-endotoxin strategies.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 8","pages":"629-35"},"PeriodicalIF":0.0,"publicationDate":"2003-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg078","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22571232","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 19

Starch-binding domain shuffling in Aspergillus niger glucoamylase. 黑曲霉葡萄糖淀粉酶中淀粉结合结构域的改组。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg066

Catherine A G Cornett, Tsuei-Yun Fang, Peter J Reilly, Clark Ford

{"title":"Starch-binding domain shuffling in Aspergillus niger glucoamylase.","authors":"Catherine A G Cornett, Tsuei-Yun Fang, Peter J Reilly, Clark Ford","doi":"10.1093/protein/gzg066","DOIUrl":"https://doi.org/10.1093/protein/gzg066","url":null,"abstract":"Aspergillus niger glucoamylase (GA) consists mainly of two forms, GAI [from the N-terminus, catalytic domain + linker + starch-binding domain (SBD)] and GAII (catalytic domain + linker). These domains were shuffled to make RGAI (SBD + linker + catalytic domain), RGAIDeltaL (SBD + catalytic domain) and RGAII (linker + catalytic domain), with domains defined by function rather than by tertiary structure. In addition, Paenibacillus macerans cyclomaltodextrin glucanotransferase SBD replaced the closely related A.niger GA SBD to give GAE. Soluble starch hydrolysis rates decreased as RGAII approximately GAII approximately GAI > RGAIDeltaL approximately RGAI approximately GAE. Insoluble starch hydrolysis rates were GAI > RGAIDeltaL > RGAI >> GAE approximately RGAII > GAII, while insoluble starch-binding capacities were GAI > RGAI > RGAIDeltaL > RGAII > GAII > GAE. These results indicate that: (i) moving the SBD to the N-terminus or replacing the native SBD somewhat affects soluble starch hydrolysis; (ii) SBD location significantly affects insoluble starch binding and hydrolysis; (iii) insoluble starch hydrolysis is imperfectly correlated with its binding by the SBD; and (iv) placing the P.macerans cyclomaltodextrin glucanotransferase SBD at the end of a linker, instead of closely associated with the rest of the enzyme, severely reduces its ability to bind and hydrolyze insoluble starch.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 7","pages":"521-9"},"PeriodicalIF":0.0,"publicationDate":"2003-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg066","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22527617","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 18

Alpha-amylase from Bacillus licheniformis mutants near to the catalytic site: effects on hydrolytic and transglycosylation activity. 靠近催化位点的地衣芽孢杆菌突变体α -淀粉酶:对水解和转糖基化活性的影响。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg060

Manuel Heriberto Rivera, Agustín López-Munguía, Xavier Soberón, Gloria Saab-Rincón

{"title":"Alpha-amylase from Bacillus licheniformis mutants near to the catalytic site: effects on hydrolytic and transglycosylation activity.","authors":"Manuel Heriberto Rivera, Agustín López-Munguía, Xavier Soberón, Gloria Saab-Rincón","doi":"10.1093/protein/gzg060","DOIUrl":"https://doi.org/10.1093/protein/gzg060","url":null,"abstract":"The alpha-amylase from Bacillus licheniformis is the most widely used enzyme in the starch industry owing to its hyperthermostability, converting starch to medium-sized oligosaccharides. Based on sequence alignment of homologous amylases, we found a semi-conserved sequence pattern near the active site between transglycosidic and hydrolytic amylases, which suggested that hydrophobicity may play a role in modifying the transglycosylation/hydrolysis ratio. Based on this analysis, we replaced residue Val286 by Phe and Tyr in Bacillus licheniformis alpha-amylase. Surprisingly, the two resultant mutant enzymes, Val286Phe and Val286Tyr, showed two different behaviors. Val286Tyr mutant was 5-fold more active for hydrolysis of starch than the wild-type enzyme. In contrast, the Val286Phe mutant, differing only by one hydroxyl group, was 3-fold less hydrolytic than the wild-type enzyme and apparently had a higher transglycosylation/hydrolysis ratio. These results are discussed in terms of affinity of subsites, hydrophobicity and electrostatic environment in the active site. The engineered enzyme reported here may represent an attractive alternative for the starch transformation industries as it affords direct and substantial material savings and requires no process modifications.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 7","pages":"505-14"},"PeriodicalIF":0.0,"publicationDate":"2003-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg060","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22528196","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 44

A long insertion reverts the functional effect of a substitution in acetylcholinesterase. 长插入恢复乙酰胆碱酯酶取代的功能效果。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg062

F Villatte, H Schulze, R D Schmid, T T Bachmann

引用次数: 3

In silico mutations and molecular dynamics studies on a winged bean chymotrypsin inhibitor protein. 飞豆胰凝乳酶抑制剂蛋白的基因突变及分子动力学研究。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg070

Jhimli Dasgupta, Udayaditya Sen, J K Dattagupta

{"title":"In silico mutations and molecular dynamics studies on a winged bean chymotrypsin inhibitor protein.","authors":"Jhimli Dasgupta, Udayaditya Sen, J K Dattagupta","doi":"10.1093/protein/gzg070","DOIUrl":"https://doi.org/10.1093/protein/gzg070","url":null,"abstract":"Winged bean chymotrypsin inhibitor (WCI) has an intruding residue Asn14 that plays a crucial role in stabilizing the reactive site loop conformation. This residue is found to be conserved in the Kunitz (STI) family of serine protease inhibitors. To understand the contribution of this scaffolding residue on the stability of the reactive site loop, it was mutated in silico to Gly, Ala, Ser, Thr, Leu and Val and molecular dynamics (MD) simulations were carried out on the mutants. The results of MD simulations reveal the conformational variability and range of motions possible for the reactive site loop of different mutants. The N-terminus side of the scissile bond, which is close to a beta-barrel, is conformationally less variable, while the C-terminus side, which is relatively far from any such secondary structural element, is more variable and needs stability through hydrogen-bonding interactions. The simulated structures of WCI and the mutants were docked in the peptide-binding groove of the cognate enzyme chymotrypsin and the ability to form standard hydrogen-bonding interactions at P3, P1 and P2' residues were compared. The results of the MD simulations coupled with docking studies indicate that hydrophobic residues like Leu and Val at the 14th position are disruptive for the integrity of the reactive site loop, whereas a residue like Thr, which can stabilize the C-terminus side of the scissile bond, can be predicted at this position. However, the size and charge of the Asn residue made it most suitable for the best maintenance of the integrity of the reactive site loop, explaining its conserved nature in the family.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 7","pages":"489-96"},"PeriodicalIF":0.0,"publicationDate":"2003-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg070","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22528194","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 16

Identification of transmembrane protein functions by binary topology patterns. 二元拓扑模式鉴定跨膜蛋白功能。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg068

Yoshiaki Sugiyama, Natalia Polulyakh, Toshio Shimizu

引用次数: 21

Differential effects of zinc on functionally distinct human growth hormone mutations. 锌对功能不同的人类生长激素突变的不同影响。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg067

K M Duda, C L Brooks

{"title":"Differential effects of zinc on functionally distinct human growth hormone mutations.","authors":"K M Duda, C L Brooks","doi":"10.1093/protein/gzg067","DOIUrl":"https://doi.org/10.1093/protein/gzg067","url":null,"abstract":"Human growth hormone (hGH) binds and activates lactogenic receptors by a sequential receptor dimerization mechanism. The affinity for the first lactogenic receptor is increased due to one zinc molecule linking hGH residues H18 and E174, located in helices 1 and 4, respectively, with two adjacent residues in the lactogenic receptor (D187 and H188). Two functionally unique groups of mutant hGHs have been identified. Addition of 25 microM zinc to lactogenic bioassays differentially affects mutant activities based on which group they belong to. One mutation (G120R) is located within the binding surface of hGH that interacts with the second lactogenic receptor. In the presence of endogenous zinc, G120R reduces the maximal activity of hGH without altering either the agonist or antagonist phases of the bell-shaped dose-response curve. Addition of zinc to this assay further reduces the activity of this protein. In contrast, mutations within a hydrophobic motif in hGH that functionally couples the two lactogenic receptor binding sites decrease the sensitivity (right-shift) of the agonist phase of the dose-response curve without similarly affecting the antagonist phase. The addition of zinc to these lactogenic assays increases the sensitivity (left-shifts) of the dose-response curves, largely negating the effect of these mutations. The effects of zinc differentiate between mutations within these two distinct functional motifs by limiting the pool of potential conformations that are available for binding within either of the receptor binding sites of this ligand.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 7","pages":"531-4"},"PeriodicalIF":0.0,"publicationDate":"2003-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg067","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22527618","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 15

Cold adaptation of a psychrophilic chitinase: a mutagenesis study. 嗜冷几丁质酶的冷适应性:诱变研究。

Protein engineering Pub Date : 2003-07-01 DOI: 10.1093/protein/gzg069

K Mavromatis, G Feller, M Kokkinidis, V Bouriotis

{"title":"Cold adaptation of a psychrophilic chitinase: a mutagenesis study.","authors":"K Mavromatis, G Feller, M Kokkinidis, V Bouriotis","doi":"10.1093/protein/gzg069","DOIUrl":"https://doi.org/10.1093/protein/gzg069","url":null,"abstract":"The gene encoding chitinase ArChiB from the Antarctic Arthrobacter sp. TAD20 has been expressed in Escherichia coli and the recombinant enzyme purified to homogeneity. In an effort to engineer cold-adapted biocatalysts through rational redesign to operate at elevated temperatures, we performed several mutations aiming to increase the rigidity of the molecular edifice of the selected psychrophilic chitinase. The mutations were designed on the basis of a homology-based three-dimensional model of the enzyme, and included an attempt to introduce a salt bridge (mutant N198K) and replacements of selected Gly residues by either Pro (mutants G93P, G254P) or Gln (G406Q). Mutant N198K resulted in a more stable protein (DeltaTm = 0.6 degrees C). Mutant G93P exhibited a DeltaTm of 1.2 degrees C, while mutants G254P and G406Q exhibited decreased stability. We conclude that the effect of mutating Gly residues on enzyme stability is rather complex and can only be understood in the context of the structural environment. Kinetic and spectroscopic analysis of these enzyme variants revealed that the kinetic parameters kcat and Km have been significantly modified.","PeriodicalId":20902,"journal":{"name":"Protein engineering","volume":"16 7","pages":"497-503"},"PeriodicalIF":0.0,"publicationDate":"2003-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/protein/gzg069","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"22528195","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 27