BMC Biochemistry最新文献

{"title":"Use of a special Brazilian red-light emitting railroad worm Luciferase in bioassays of NEK7 protein Kinase and Creatine Kinase.","authors":"Arina Marina Perez,&nbsp;Bruno Aquino,&nbsp;Vadim Viviani,&nbsp;Jörg Kobarg","doi":"10.1186/s12858-017-0087-z","DOIUrl":"https://doi.org/10.1186/s12858-017-0087-z","url":null,"abstract":"<p><strong>Background: </strong>Luciferases, enzymes that catalyze bioluminescent reactions in different organisms, have been extensively used for bioanalytical purposes. The most well studied bioluminescent system is that of firefly and other beetles, which depends on a luciferase, a benzothiazolic luciferin and ATP, and it is being widely used as a bioanalytical reagent to quantify ATP. Protein kinases are proteins that modify other proteins by transferring phosphate groups from a nucleoside triphosphate, usually ATP.</p><p><strong>Methods: </strong>Here, we used a red-light emitting luciferase from Phrixotrix hirtus railroad worm to determine the activity of kinases in a coupled assay, based on luminescence that is generated when luciferase is in the presence of its substrate, the luciferin, and ATP.</p><p><strong>Results: </strong>In this work we used, after several optimization reactions, creatine kinase isoforms as well as NEK7 protein kinase in the absence or presence of ATP analogous inhibitors  to validate this new luminescence method.</p><p><strong>Conclusion: </strong>With this new approach we validated a luminescence method to quantify kinase activity, with different substrates and inhibition screening tests, using a novel red-light emitting luciferase as a reporter enzyme.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"12"},"PeriodicalIF":0.0,"publicationDate":"2017-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0087-z","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35182215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dacin, one metalloproteinase from Deinagkistrodon acutus venom inhibiting contraction of mouse ileum muscle.","authors":"Bin Zhou,&nbsp;Gang Liu,&nbsp;Qiyi He,&nbsp;Bo Li,&nbsp;Xiaodong Yu","doi":"10.1186/s12858-017-0086-0","DOIUrl":"https://doi.org/10.1186/s12858-017-0086-0","url":null,"abstract":"<p><strong>Background: </strong>Mice were bitten by five-pace vipers (Deinagkistrodon acutus), and then envenomed. It was well-known that the snake venom mainly disturbed the blood homeostasis of the envenomed victims. Ocassionally, we found that the venom of D. acutus could inhibit the contraction tension of mouse ileum, so in this study we aimed to identify the active component inhibiting the contraction tension of mouse ileum in the snake venom.</p><p><strong>Results: </strong>The active component inhibiting the contraction tension of mouse ileum, designated as Dacin, was isolated from D. acutus venom, purified to protein homogeneity and composed of a single peptide chain, about 23 kDa analyzed by SDS-PAGE, and 22, 947. 9 Da measured by MALDI-TOF-MS. Not only the results of its PMF blasted by Mascot indicated that Dacin may be one snake venom metalloproteinase (SVMP), but also the results of the biochemical and in-vivo assays as follow demonstrated that it was one SVMP: it cleaved Aα and Bβ chains, not Cγ of bovine fibrinogen within 1 h, and also hydrolyzed fibrin polymer; besides its fibrino(geno)lytic activities were strongly inhibited by β- mercaptoethanol, EDTA and EGTA; and it could induce a hemorrhagic reaction under the dorsal skin of mouse. In the isolated tissue assays, Dacin caused the concentration-dependent and time-dependent inhibitory actions on the spontaneous contraction tension of the ileum smooth muscle of mouse, and the inhibitory effects were irreversible.</p><p><strong>Conclusions: </strong>Taken together, for the first time one active component (Dacin, a SVMP) that irreversibly inhibited the spontaneous contraction tension of mouse ileum has been isolated and identified from D. acutus venom. The findings may provide not only a new insight for toxicological researches on SVMPs and venoms of the vipers, but also a reference for clinicians to treat the snake-bitten victims. However, Dacin's inhibitory molecular mechanism will be further studied in the future.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"11"},"PeriodicalIF":0.0,"publicationDate":"2017-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0086-0","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35161784","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Serendipitous discovery of light-induced (In Situ) formation of an Azo-bridged dimeric sulfonated naphthol as a potent PTP1B inhibitor.","authors":"Robert D Bongard, Michael Lepley, Khushabu Thakur, Marat R Talipov, Jaladhi Nayak, Rachel A Jones Lipinski, Chris Bohl, Noreena Sweeney, Ramani Ramchandran, Rajendra Rathore, Daniel S Sem","doi":"10.1186/s12858-017-0083-3","DOIUrl":"10.1186/s12858-017-0083-3","url":null,"abstract":"<p><strong>Background: </strong>Protein tyrosine phosphatases (PTPs) like dual specificity phosphatase 5 (DUSP5) and protein tyrosine phosphatase 1B (PTP1B) are drug targets for diseases that include cancer, diabetes, and vascular disorders such as hemangiomas. The PTPs are also known to be notoriously difficult targets for designing inihibitors that become viable drug leads. Therefore, the pipeline for approved drugs in this class is minimal. Furthermore, drug screening for targets like PTPs often produce false positive and false negative results.</p><p><strong>Results: </strong>Studies presented herein provide important insights into: (a) how to detect such artifacts, (b) the importance of compound re-synthesis and verification, and (c) how in situ chemical reactivity of compounds, when diagnosed and characterized, can actually lead to serendipitous discovery of valuable new lead molecules. Initial docking of compounds from the National Cancer Institute (NCI), followed by experimental testing in enzyme inhibition assays, identified an inhibitor of DUSP5. Subsequent control experiments revealed that this compound demonstrated time-dependent inhibition, and also a time-dependent change in color of the inhibitor that correlated with potency of inhibition. In addition, the compound activity varied depending on vendor source. We hypothesized, and then confirmed by synthesis of the compound, that the actual inhibitor of DUSP5 was a dimeric form of the original inhibitor compound, formed upon exposure to light and oxygen. This compound has an IC₅₀ of 36 μM for DUSP5, and is a competitive inhibitor. Testing against PTP1B, for selectivity, demonstrated the dimeric compound was actually a more potent inhibitor of PTP1B, with an IC₅₀ of 2.1 μM. The compound, an azo-bridged dimer of sulfonated naphthol rings, resembles previously reported PTP inhibitors, but with 18-fold selectivity for PTP1B versus DUSP5.</p><p><strong>Conclusion: </strong>We report the identification of a potent PTP1B inhibitor that was initially identified in a screen for DUSP5, implying common mechanism of inhibitory action for these scaffolds.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"10"},"PeriodicalIF":0.0,"publicationDate":"2017-05-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5452347/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35050978","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Chemical and structural characterization of α-N-acetylgalactosaminidase I and II from starfish, asterina amurensis.","authors":"Md Harun-Or Rashid,&nbsp;Golam Sadik,&nbsp;Ahm Khurshid Alam,&nbsp;Toshihisa Tanaka","doi":"10.1186/s12858-017-0085-1","DOIUrl":"https://doi.org/10.1186/s12858-017-0085-1","url":null,"abstract":"<p><strong>Background: </strong>The marine invertebrate starfish was found to contain a novel α-N-acetylgalactosaminidase, α-GalNAcase II, which catalyzes removal of terminal α-N-acetylgalactosamine (α-GalNAc), in addition to a typical α-N-acetylgalactosaminidase, α-GalNAcase I, which catalyzes removal of terminal α-N-acetylgalactosamine (α-GalNAc) and, to a lesser extent, galactose. The interrelationship between α-GalNAcase I and α-GalNAcase II and the molecular basis of their differences in substrate specificity remain unknown.</p><p><strong>Results: </strong>Chemical and structural comparisons between α-GalNAcase I and II using immunostaining, N-terminal amino acid sequencing and peptide analysis showed high homology to each other and also to other glycoside hydrolase family (GHF) 27 members. The amino acid sequence of peptides showed conserved residues at the active site as seen in typical α-GalNAcase. Some substitutions of conserved amino acid residues were found in α-GalNAcase II that were located near catalytic site. Among them G171 and A173, in place of C171 and W173, respectively in α-GalNAcase were identified to be responsible for lacking intrinsic α-galactosidase activity of α-GalNAcase II. Chemical modifications supported the presence of serine, aspartate and tryptophan as active site residues. Two tryptophan residues (W16 and W173) were involved in α-galactosidase activity, and one (W16) of them was involved in α-GalNAcase activity.</p><p><strong>Conclusions: </strong>The results suggested that α-GalNAcase I and II are closely related with respect to primary and higher order structure and that their structural differences are responsible for difference in substrate specificities.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"9"},"PeriodicalIF":0.0,"publicationDate":"2017-05-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0085-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35028752","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural insight into the inactivation of Mycobacterium tuberculosis non-classical transpeptidase Ldt_Mt2 by biapenem and tebipenem.","authors":"Mario A Bianchet,&nbsp;Ying H Pan,&nbsp;Leighanne A Brammer Basta,&nbsp;Harry Saavedra,&nbsp;Evan P Lloyd,&nbsp;Pankaj Kumar,&nbsp;Rohini Mattoo,&nbsp;Craig A Townsend,&nbsp;Gyanu Lamichhane","doi":"10.1186/s12858-017-0082-4","DOIUrl":"https://doi.org/10.1186/s12858-017-0082-4","url":null,"abstract":"<p><strong>Background: </strong>The carbapenem subclass of β-lactams is among the most potent antibiotics available today. Emerging evidence shows that, unlike other subclasses of β-lactams, carbapenems bind to and inhibit non-classical transpeptidases (L,D-transpeptidases) that generate 3 → 3 linkages in bacterial peptidoglycan. The carbapenems biapenem and tebipenem exhibit therapeutically valuable potencies against Mycobacterium tuberculosis (Mtb).</p><p><strong>Results: </strong>Here, we report the X-ray crystal structures of Mtb L,D-transpeptidase-2 (Ldt_Mt2) complexed with biapenem or tebipenem. Despite significant variations in carbapenem sulfur side chains, biapenem and tebipenem ultimately form an identical adduct that docks to the outer cavity of Ldt_Mt2. We propose that this common adduct is an enzyme catalyzed decomposition of the carbapenem adduct by a mechanism similar to S-conjugate elimination by β-lyases.</p><p><strong>Conclusion: </strong>The results presented here demonstrate biapenem and tebipenem bind to the outer cavity of Ldt_Mt2, covalently inactivate the enzyme, and subsequently degrade via an S-conjugate elimination mechanism. We discuss structure based drug design based on the findings and propose that the S-conjugate elimination can be leveraged to design novel agents to deliver and locally release antimicrobial factors to act synergistically with the carbapenem carrier.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"8"},"PeriodicalIF":0.0,"publicationDate":"2017-05-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0082-4","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35028754","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CrMAPK3 regulates the expression of iron-deficiency-responsive genes in Chlamydomonas reinhardtii.","authors":"Xiaowen Fei,&nbsp;Junmei Yu,&nbsp;Yajun Li,&nbsp;Xiaodong Deng","doi":"10.1186/s12858-017-0081-5","DOIUrl":"https://doi.org/10.1186/s12858-017-0081-5","url":null,"abstract":"<p><strong>Background: </strong>Under iron-deficient conditions, Chlamydomonas exhibits high affinity for iron absorption. Nevertheless, the response, transmission, and regulation of downstream gene expression in algae cells have not to be investigated. Considering that the MAPK pathway is essential for abiotic stress responses, we determined whether this pathway is involved in iron deficiency signal transduction in Chlamydomonas.</p><p><strong>Results: </strong>Arabidopsis MAPK gene sequences were used as entry data to search for homologous genes in Chlamydomonas reinhardtii genome database to investigate the functions of mitogen-activated protein kinase (MAPK) gene family in C. reinhardtii under iron-free conditions. Results revealed 16 C. reinhardtii MAPK genes labeled CrMAPK2-CrMAPK17 with TXY conserved domains and low homology to MAPK in yeast, Arabidopsis, and humans. The expression levels of these genes were then analyzed through qRT-PCR and exposure to high salt (150 mM NaCl), low nitrogen, or iron-free conditions. The expression levels of these genes were also subjected to adverse stress conditions. The mRNA levels of CrMAPK2, CrMAPK3, CrMAPK4, CrMAPK5, CrMAPK6, CrMAPK8, CrMAPK9, and CrMAPK11 were remarkably upregulated under iron-deficient stress. The increase in CrMAPK3 expression was 43-fold greater than that in the control. An RNA interference vector was constructed and transformed into C. reinhardtii 2A38, an algal strain with an exogenous FOX1:ARS chimeric gene, to silence CrMAPK3. After this gene was silenced, the mRNA levels and ARS activities of FOX1:ARS chimeric gene and endogenous CrFOX1 were decreased. The mRNA levels of iron-responsive genes, such as CrNRAMP2, CrATX1, CrFTR1, and CrFEA1, were also remarkably reduced.</p><p><strong>Conclusion: </strong>CrMAPK3 regulates the expression of iron-deficiency-responsive genes in C. reinhardtii.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"6"},"PeriodicalIF":0.0,"publicationDate":"2017-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0081-5","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35001432","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural similarities and functional differences clarify evolutionary relationships between tRNA healing enzymes and the myelin enzyme CNPase.","authors":"Gopinath Muruganandam, Arne Raasakka, Matti Myllykoski, Inari Kursula, Petri Kursula","doi":"10.1186/s12858-017-0084-2","DOIUrl":"10.1186/s12858-017-0084-2","url":null,"abstract":"<p><strong>Background: </strong>Eukaryotic tRNA splicing is an essential process in the transformation of a primary tRNA transcript into a mature functional tRNA molecule. 5'-phosphate ligation involves two steps: a healing reaction catalyzed by polynucleotide kinase (PNK) in association with cyclic phosphodiesterase (CPDase), and a sealing reaction catalyzed by an RNA ligase. The enzymes that catalyze tRNA healing in yeast and higher eukaryotes are homologous to the members of the 2H phosphoesterase superfamily, in particular to the vertebrate myelin enzyme 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase).</p><p><strong>Results: </strong>We employed different biophysical and biochemical methods to elucidate the overall structural and functional features of the tRNA healing enzymes yeast Trl1 PNK/CPDase and lancelet PNK/CPDase and compared them with vertebrate CNPase. The yeast and the lancelet enzymes have cyclic phosphodiesterase and polynucleotide kinase activity, while vertebrate CNPase lacks PNK activity. In addition, we also show that the healing enzymes are structurally similar to the vertebrate CNPase by applying synchrotron radiation circular dichroism spectroscopy and small-angle X-ray scattering.</p><p><strong>Conclusions: </strong>We provide a structural analysis of the tRNA healing enzyme PNK and CPDase domains together. Our results support evolution of vertebrate CNPase from tRNA healing enzymes with a loss of function at its N-terminal PNK-like domain.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"7"},"PeriodicalIF":0.0,"publicationDate":"2017-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5434554/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"35000974","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Binding of smoothelin-like 1 to tropomyosin and calmodulin is mutually exclusive and regulated by phosphorylation.","authors":"Annegret Ulke-Lemée,&nbsp;David Hao Sun,&nbsp;Hiroaki Ishida,&nbsp;Hans J Vogel,&nbsp;Justin A MacDonald","doi":"10.1186/s12858-017-0080-6","DOIUrl":"https://doi.org/10.1186/s12858-017-0080-6","url":null,"abstract":"<p><strong>Background: </strong>The smoothelin-like 1 protein (SMTNL1) can associate with tropomyosin (Tpm) and calmodulin (CaM), two proteins essential to the smooth muscle contractile process. SMTNL1 is phosphorylated at Ser301 by protein kinase A during calcium desensitization in smooth muscle, yet the effect of SMTNL1 phosphorylation on Tpm- and CaM-binding has yet to be investigated.</p><p><strong>Results: </strong>Using pull down studies with Tpm-Sepharose and CaM-Sepharose, we examined the interplay between Tpm binding, CaM binding, phosphorylation of SMTNL1 and calcium concentration. Phosphorylation greatly enhanced the ability of SMTNL1 to associate with Tpm in vitro; surface plasmon resonance yielded a 10-fold enhancement in K _D value with phosphorylation. The effect on CaM binding is more complex and varies with the availability of calcium.</p><p><strong>Conclusions: </strong>Combining both CaM and Tpm with SMTNL1 shows that the binding to both is mutually exclusive.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"5"},"PeriodicalIF":0.0,"publicationDate":"2017-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0080-6","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34835619","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Increased enzymatic hydrolysis of sugarcane bagasse by a novel glucose- and xylose-stimulated β-glucosidase from Anoxybacillus flavithermus subsp. yunnanensis E13^T.","authors":"Yang Liu,&nbsp;Rui Li,&nbsp;Jing Wang,&nbsp;Xiaohan Zhang,&nbsp;Rong Jia,&nbsp;Yi Gao,&nbsp;Hui Peng","doi":"10.1186/s12858-017-0079-z","DOIUrl":"https://doi.org/10.1186/s12858-017-0079-z","url":null,"abstract":"<p><strong>Background: </strong>β-Glucosidase is claimed as a key enzyme in cellulose hydrolysis. The cellulosic fibers are usually entrapped with hemicelluloses containing xylose. So there is ongoing interest in searching for glucose- and xylose-stimulated β-glucosidases to increase the efficiency of hydrolysis of cellulosic biomass.</p><p><strong>Results: </strong>A thermostable β-glucosidase gene (Bglp) was cloned from Anoxybacillus flavithermus subsp. yunnanensis E13^T and characterized. Optimal enzyme activity was observed at 60 °C and pH 7.0. Bglp was relatively stable at 60 °C with a 10-h half-life. The kinetic parameters V _max and K _m for p-nitrophenyl-β-D-glucopyranoside (pNPG) were 771 ± 39 μmol/min/mg and 0.29 ± 0.01 mM, respectively. The activity of Bglp is dramatically stimulated by glucose or xylose at concentrations up to 1.4 M. After Bglp was added to Celluclast® 1.5 L, the conversion of sugarcane bagasse was 48.4 ± 0.8%, which was much higher than of Celluclast® 1.5 L alone. Furthermore, Bglp showed obvious advantages in the hydrolysis when initial concentrations of glucose and xylose are high.</p><p><strong>Conclusions: </strong>The supplementation of BglP significantly enhanced the glucose yield from sugarcane bagasse, especially in the presence of high concentrations of glucose or xylose. Bglp should be a promising candidate for industrial applications.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"4"},"PeriodicalIF":0.0,"publicationDate":"2017-03-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0079-z","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34819532","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Systematic substitutions at BLIP position 50 result in changes in binding specificity for class A β-lactamases.","authors":"Carolyn J Adamski,&nbsp;Timothy Palzkill","doi":"10.1186/s12858-017-0077-1","DOIUrl":"https://doi.org/10.1186/s12858-017-0077-1","url":null,"abstract":"<p><strong>Background: </strong>The production of β-lactamases by bacteria is the most common mechanism of resistance to the widely prescribed β-lactam antibiotics. β-lactamase inhibitory protein (BLIP) competitively inhibits class A β-lactamases via two binding loops that occlude the active site. It has been shown that BLIP Tyr50 is a specificity determinant in that substitutions at this position result in large differential changes in the relative affinity of BLIP for class A β-lactamases.</p><p><strong>Results: </strong>In this study, the effect of systematic substitutions at BLIP position 50 on binding to class A β-lactamases was examined to further explore the role of BLIP Tyr50 in modulating specificity. The results indicate the sequence requirements at position 50 are widely different depending on the target β-lactamase. Stringent sequence requirements were observed at Tyr50 for binding Bacillus anthracis Bla1 while moderate requirements for binding TEM-1 and relaxed requirements for binding KPC-2 β-lactamase were seen. These findings cannot be easily rationalized based on the β-lactamase residues in direct contact with BLIP Tyr50 since they are identical for Bla1 and KPC-2 suggesting that differences in the BLIP-β-lactamase interface outside the local environment of Tyr50 influence the effect of substitutions.</p><p><strong>Conclusions: </strong>Results from this study and previous studies suggest that substitutions at BLIP Tyr50 may induce changes at the interface outside its local environment and point to the complexity of predicting the impact of substitutions at a protein-protein interaction interface.</p>","PeriodicalId":9113,"journal":{"name":"BMC Biochemistry","volume":"18 1","pages":"2"},"PeriodicalIF":0.0,"publicationDate":"2017-03-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1186/s12858-017-0077-1","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"34786433","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}