Biochimica et biophysica acta. Proteins and proteomics最新文献

{"title":"Investigation of adipocyte differentiation based on proteomics and intact N-glycopeptide modificationomics","authors":"Xin-Yu Li ,&nbsp;Nuerbiye Nuermaimaiti ,&nbsp;Xuanyu Meng ,&nbsp;Xiaozheng Zhang ,&nbsp;Aikedaimu Abudukeremu ,&nbsp;Yihuai He ,&nbsp;Wenting Ma ,&nbsp;Xuelei Chen ,&nbsp;Shangkun Li ,&nbsp;Jiaxin Sun ,&nbsp;Yaqun Guan","doi":"10.1016/j.bbapap.2024.141052","DOIUrl":"10.1016/j.bbapap.2024.141052","url":null,"abstract":"<div><h3>Objective</h3><div>To investigate the role of N-glycosylation modification of proteins in adipocyte differentiation during the adipogenic process.</div></div><div><h3>Methods</h3><div>SVF cells and adipocytes were analyzed for proteomics and intact N-glycopeptide modificationomics.Differential expression of proteins, glycoforms, and sites between the two groups was screened and subjected to Gene Ontology (GO) functional enrichment analysis, KEGG pathway enrichment analysis, and protein-protein interaction (PPI) network analysis. The top 20 most significantly differentially expressed adipogenic differentiation-related proteins were identified, and the most pronouncedly altered proteins were analyzed for glycoforms, glycan chains, and sites.</div></div><div><h3>Results</h3><div>Proteomics analysis identified 39,392 peptides and 5208 proteins, while intact N-glycopeptide modification profiling identified 3293 intact glycopeptides, 426 proteins, and 161 glycan chains. Proteomics identified 2510 differentially expressed proteins, with CD36 (Cluster of Differentiation 36, CD36) significantly upregulated. In adipocytes, CD36 had 4 N-glycosylation sites: N79, N220, N320, N417, with N320 being a newly identified site. GO enrichment results indicated that CD36 is associated with fatty acid oxidation, lipid oxidation, and fatty acid uptake into cells.</div></div><div><h3>Conclusion</h3><div>Multiple proteins undergo N-glycosylation modification during adipocyte differentiation, with CD36, a fatty acid translocase, being significantly expressed in adipocytes. This suggests that N-glycosylation modification of CD36 may play a crucial role in adipocyte differentiation, providing a foundation for further investigation into the function of CD36 N-glycosylation in adipocyte differentiation.</div></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141052"},"PeriodicalIF":2.5,"publicationDate":"2024-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142387571","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural insights into the mechanism underlying the dual cofactor specificity of glyoxylate reductase from Acetobacter aceti in the β-hydroxyacid dehydrogenase family","authors":"Toma Rani Majumder ,&nbsp;Takuya Yoshizawa ,&nbsp;Masao Inoue ,&nbsp;Riku Aono ,&nbsp;Hiroyoshi Matsumura ,&nbsp;Hisaaki Mihara","doi":"10.1016/j.bbapap.2024.141051","DOIUrl":"10.1016/j.bbapap.2024.141051","url":null,"abstract":"<div><div>The β-hydroxyacid dehydrogenase family exhibits diverse cofactor preferences: some enzymes favor NAD, others favor NADP, and a subset can utilize both NAD and NADPH. Glyoxylate reductase from <em>Acetobacter aceti</em> JCM 20276 (AacGR) exhibits a dual cofactor specificity for NADPH and NADH in its catalytic reduction of glyoxylate to glycolate. In contrast to conventional cofactor-discriminating motifs, NRX and DXX, found in NADP- and NAD-specific enzymes, respectively, AacGR has a TPS motif in the equivalent position. Here we report X-ray crystallographic analysis of AacGR in its ligand-free form, and in complexes with NADPH and NADH, revealing critical interactions: Ser41 of the TPS motif interacted with the 2′-phosphate group of NADPH, while no analogous interaction occurred with the ribose hydroxy groups of NADH. Moreover, the TPS motif resided within a characteristic β-turn-like structure adjacent to a long flexible loop. Site-directed mutagenesis and kinetic analyses suggest that Ser41 facilitates NADPH binding, while the lack of a direct interaction of the TPS motif with NADH may allow for NADH utilization. The conformational dynamics of the TPS-containing β-turn-like structure along with the flexible loop likely govern the dual cofactor specificity and catalytic turnover of AacGR.</div></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141051"},"PeriodicalIF":2.5,"publicationDate":"2024-10-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142379056","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structure and functional studies of Avt1, a novel peptide from the sea anemone Aulactinia veratra","authors":"Renad A. Albar ,&nbsp;Hayden L. Smith ,&nbsp;Karoline Sanches ,&nbsp;Dorothy C.C. Wai ,&nbsp;Muhammad Umair Naseem ,&nbsp;Tibor G. Szanto ,&nbsp;Gyorgy Panyi ,&nbsp;Peter J. Prentis ,&nbsp;Raymond S. Norton","doi":"10.1016/j.bbapap.2024.141050","DOIUrl":"10.1016/j.bbapap.2024.141050","url":null,"abstract":"<div><div>Sea anemones are a rich source of peptide toxins spanning a diverse range of biological activities, typically targeting proteins such as ion channels, receptors and transporters. These peptide toxins and their analogues are usually highly stable and selective for their molecular targets, rendering them of interest as molecular tools, insecticides and therapeutics. Recent transcriptomic and proteomic analyses of the sea anemone <em>Aulactinia veratra</em> identified a novel 28-residue peptide, designated Avt1. Avt1 was produced using solid-phase peptide synthesis, followed by oxidative folding and purification of the folded peptide using reversed-phase high-performance liquid chromatography. The liquid chromatography-mass spectrometry profile of synthetic Avt1 showed a pure peak with molecular mass 6 Da less than that of the reduced form of the peptide, indicating the successful formation of three disulfide bonds. The solution structure determined by NMR revealed that Avt1 adopts an inhibitor cystine knot (ICK) fold, in which a ring is formed by two disulfide bonds with a third disulfide penetrating the ring to create the pseudo-knot. This structure provides ICK peptides with high structural, thermal and proteolytic stability. Consistent with its ICK structure, Avt1 was resistant to proteolysis by trypsin, chymotrypsin and pepsin, although it was not a trypsin inhibitor. Avt1 at 100 nM showed no activity in patch–clamp electrophysiological assays against several mammalian voltage-gated ion channels, but has structural features similar to toxins targeting insect sodium ion channels. Although sequence homologues of Avt1 are found in a number of sea anemones, this is the first representative of this family to be characterised structurally and functionally.</div></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141050"},"PeriodicalIF":2.5,"publicationDate":"2024-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142364212","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of dystrophin Dp71dΔ71-associated proteins in PC12 cells by quantitative proteomics","authors":"Coztli Azotla-Vilchis ,&nbsp;Candelaria Merino-Jiménez ,&nbsp;Emmanuel Ríos-Castro ,&nbsp;Jorge Aragón ,&nbsp;Víctor Ceja ,&nbsp;Cecilia Montanez","doi":"10.1016/j.bbapap.2024.141049","DOIUrl":"10.1016/j.bbapap.2024.141049","url":null,"abstract":"<div><div>Dystrophin Dp71 is essential for the development of the nervous system. Its alteration is associated with intellectual disability. Different Dp71 isoforms are generated by alternative splicing; however, their functions have not been fully described. Here, we identified Dp71d_Δ71-associated proteins to understand the complex functions. PC12 cells, stably transfected with pTRE2pur-Myc/Dp71d_Δ71 or pTRE2pur-Myc empty vector (EV), were analyzed by immunoprecipitation followed with quantitative proteomics with data-independent acquisition and ion mobility separation. We used the Top3 method to quantify absolutely every protein detected. A total of 106 proteins were quantified with Progenesis QI software and the database UP000002494. Seven new proteins associated with Dp71d_Δ71 were selected with at least 2-fold quantity between immunoprecipitated proteins of PC12-Myc/Dp71d_Δ71 versus PC12-EV cells. These results revealed new proteins that interact with Dp71d_Δ71, including β-Tubulin, S-adenosylmethionine synthase isoform type-2, adapter molecule crk, helicase with zinc finger 2, WD repeat domain 93, cyclin-L2 and myosin-10, which are related to cell migration and/or cell growth. The results lay the foundation for future research on the relationship between these proteins and Dp71 isoforms.</div></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141049"},"PeriodicalIF":2.5,"publicationDate":"2024-09-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142340302","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Assembly of Hydrophobin class I from Agaricus bisporus produced different amyloid-like fibrils","authors":"Jesús Rojas-Osnaya,&nbsp;Hugo Nájera","doi":"10.1016/j.bbapap.2024.141048","DOIUrl":"10.1016/j.bbapap.2024.141048","url":null,"abstract":"<div><div>This work studied the extraction, purification, characterization, and assembly of hydrophobin class I from <em>Agaricus bisporus</em> (ABH4)<em>.</em> The highest soluble protein concentration was obtained from the pinhead, the extraction and purification were efficient for hydrophobin class I, obtaining a band of 12 kDa. The identified sequence of hydrophobin presented the eight cysteine residues; for the prediction of the structure, hydrophobin presented more alpha helix structures than beta sheets. It was observed that the hydrophobin managed to decrease and increase the contact angle in Teflon and glass, respectively, finding a micellar critical concentration of 221 μg mL⁻¹. ThT experiments demonstrated that the production of fibrils decreased at basic pH, while acidic and neutral pH favoured the formation of fibrils. Likewise, the addition of colloidal Teflon affects the formation of fibrils. Circular dichroism spectra proved that hydrophobin class I undergo changes in its secondary structure, increasing its alpha helix and beta sheet content after vortexing. It was observed that the analysis by scanning electron microscopy and atomic force microscopy of the hydrophobin produced different amyloid-like structures in glass and mica.</div></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141048"},"PeriodicalIF":2.5,"publicationDate":"2024-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142340301","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Serratiopeptidase exhibits antibiofilm activity through the proteolytic function of N-terminal domain and versatile function of the C-terminal domain","authors":"Vishal Srivastava,&nbsp;Sheetal Bandhu,&nbsp;Shivam Mishra,&nbsp;Tapan K. Chaudhuri","doi":"10.1016/j.bbapap.2024.141046","DOIUrl":"10.1016/j.bbapap.2024.141046","url":null,"abstract":"<div><h3>Background</h3><p>Serratiopeptidase, a serine protease traditionally used as an oral anti-inflammatory drug has been found to show antibiofilm action. Structurally, it comprises of two distinct domains; viz-the N-terminal catalytic domain (N<em>cat</em>) and a C-terminal RTX (Repeat-In-Toxin) domain (C<em>rtx</em>). Understanding the antibiofilm action of the serratiopeptidase molecule, as well as the antibiofilm action of each of its two domains, was the objective of this study.</p></div><div><h3>Results</h3><p>Separate clones to express the complete recombinant serratiopeptidase protein and its variant containing a mutation in the catalytic site, the N-terminal catalytic domain and its mutant, and the C-terminal Repeat-In-Toxin domain were prepared, and the proteins were purified. The impact of these proteins on pre-existing biofilms, as well as their effect upon addition of these proteins during biofilm formation was investigated.</p></div><div><h3>Conclusions</h3><p>In our investigation, we have been able to analyze the antibiofilm action of serratiopeptidase in detail. Obtained results conclude that while N-terminally located proteolytic domain of serratiopeptidase conventionally acts against biofilms by hydrolytic activity, the C-terminal domain regulates or prevents biofilm formation by yet unknown mechanism in addition to its known function as an C-terminal located calcium modulated internal chaperone ensuring the proper folding and secretion of the molecule. The study's findings give new evidence that the C<em>rtx</em> domain plays a significant role in antibiofilm action. The proteolytic N<em>cat</em> domain breaks down pre-formed biofilms. The C-terminal domain, on the other hand, acts as an inhibitor of biofilm formation by regulating or preventing biofilm development.</p></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141046"},"PeriodicalIF":2.5,"publicationDate":"2024-09-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S1570963924000530/pdfft?md5=3a4d2c38465f942f4b2f8b47b6076387&pid=1-s2.0-S1570963924000530-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142145008","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"From sequence to function: Exploring biophysical properties of bacteriophage BFK20 lytic transglycosylase domain from the minor tail protein gp15","authors":"Kristina Papayova ,&nbsp;Lucia Bocanova ,&nbsp;Vladena Bauerova ,&nbsp;Jacob Bauer ,&nbsp;Nora Halgasova ,&nbsp;Maria Kajsikova ,&nbsp;Gabriela Bukovska","doi":"10.1016/j.bbapap.2024.141044","DOIUrl":"10.1016/j.bbapap.2024.141044","url":null,"abstract":"<div><p>Bacteriophages have evolved different mechanisms of infection and penetration of bacterial cell walls. In <em>Siphoviridae</em>-like viruses, the inner tail proteins have a pivotal role in these processes and often encode lytic protein domains which increase infection efficiency. A soluble lytic transglycosylase (SLT) domain was identified in the minor tail protein gp15 from the BFK20 bacteriophage. Six fragments containing this SLT domain with adjacent regions of different lengths were cloned, expressed and purified. The biophysical properties of the two best expressing fragments were characterized by nanoDSF and CD spectroscopy, which showed that both fragments had a high refolding ability of 90 %. 3D modeling indicated that the bacteriophage BFK20 SLT domain is structurally similar to lysozyme. The degradation activity of these SLT proteins was evaluated using a lysozyme activity assay. BFK20 might use its transglycosylase activity to allow efficient phage DNA entry into the host cell by degrading bacterial peptidoglycan.</p></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1873 1","pages":"Article 141044"},"PeriodicalIF":2.5,"publicationDate":"2024-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142103945","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Assigning roles in Chlamydomonas ribosome biogenesis: The conserved factor NIP7","authors":"Raissa Ferreira Gutierrez ,&nbsp;Heloisa Ciol ,&nbsp;Angélica L. Carrillo Barra ,&nbsp;Diego Antonio Leonardo ,&nbsp;Juliana S. Avaca-Crusca ,&nbsp;Otavio H. Thiemann ,&nbsp;Nilson Ivo Tonin Zanchin ,&nbsp;Ana P. Ulian Araujo","doi":"10.1016/j.bbapap.2024.141045","DOIUrl":"10.1016/j.bbapap.2024.141045","url":null,"abstract":"<div><p>Ribosome biogenesis (RB) is a highly conserved process across eukaryotes that results in the assembly of functional ribosomal subunits. Studies in <em>Saccharomyces cerevisiae</em> and <em>Homo sapiens</em> have identified numerous RB factors (RBFs), including the NIP7 protein, which is involved in late-stage pre-60S ribosomal maturation. NIP7 expression has also been observed in <em>Chlamydomonas reinhardtii</em>, highlighting its evolutionary significance. This study aimed to characterize the function of the NIP7 protein from <em>C. reinhardtii</em> (<em>Cr</em>Nip7) through protein complementation assays and a paromomycin resistance test, assessing its ability to complement the role of NIP7 in yeast. Protein interaction studies were conducted via yeast two-hybrid assay to identify potential protein partners of <em>Cr</em>Nip7. Additionally, rRNA modeling analysis was performed using the predicted structure of <em>Cr</em>Nip7 to investigate its interaction with rRNA. The study revealed that <em>Cr</em>Nip7 can complement the role of NIP7 in yeast, implicating <em>Cr</em>Nip7 in the biogenesis of the 60S ribosomal subunit. Furthermore, two possible partner proteins of CrNip7, UNC-p and G-patch, were identified through yeast two-hybrid assay. The potential of these proteins to interact with CrNip7 was explored through in silico analyses. Furthermore, nucleic acid interaction was also evaluated, indicating the involvement of the N- and C-terminal domains of CrNIP7 in interacting with rRNA. Collectively, our findings provide valuable insights into the RBFs CrNip7, offering novel information for comparative studies on RB among eukaryotic model organisms, shedding light on its evolutionary conservation and functional role across species.</p></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1872 6","pages":"Article 141045"},"PeriodicalIF":2.5,"publicationDate":"2024-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142103919","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Identification of potential pharmacological chaperones that selectively stabilize mutated Aspartoacylases in Canavan disease","authors":"Nitesh Kumar Poddar ,&nbsp;Yasanandana S. Wijayasinghe ,&nbsp;Ronald E. Viola","doi":"10.1016/j.bbapap.2024.141043","DOIUrl":"10.1016/j.bbapap.2024.141043","url":null,"abstract":"<div><p>Canavan disease is caused by mutations in the <em>ASPA</em> gene, leading to diminished catalytic activity of aspartoacylase in the brain. Clinical missense mutations are found throughout the enzyme structure, with many of these mutated enzymes having not only decreased activity but also compromised stability. High-throughput screening of a small molecule library has identified several compounds that significantly increase the thermal stability of the E285A mutant enzyme, the most predominant clinical mutation in Canavan disease, while having a negligible effect on the native enzyme. Based on the initial successes, some structural analogs of these initial hits were selected for further examination. Glutathione, NAAG and patulin were each confirmed to be competitive inhibitors, indicating the binding of these compounds at the dimer interface or near the active site of the E285A enzyme. The experimental results were theoretically examined with the help of the docking analysis method. The structure activity-guided optimization of these compounds can potentially lead to potential pharmacological chaperones that could alleviate the detrimental effect of <em>ASPA</em> mutations in Canavan patients.</p></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1872 6","pages":"Article 141043"},"PeriodicalIF":2.5,"publicationDate":"2024-08-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141916040","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Kinetic characterization of the C-terminal domain of Malonyl-CoA reductase","authors":"Mirela Tkalcic Cavuzic (Tkalčić Čavužić) ,&nbsp;Amanda Silva de Sousa ,&nbsp;Jeremy R. Lohman ,&nbsp;Grover L. Waldrop","doi":"10.1016/j.bbapap.2024.141033","DOIUrl":"10.1016/j.bbapap.2024.141033","url":null,"abstract":"<div><p>Malonyl-CoA reductase utilizes two equivalents of NADPH to catalyze the reduction of malonyl-CoA to 3-hydroxypropionic acid (3HP). This reaction is part of the carbon fixation pathway in the phototrophic bacterium <em>Chloroflexus aurantiacus</em>. The enzyme is composed of two domains. The C-terminal domain catalyzes the reduction of malonyl-CoA to malonic semialdehyde, while the N-terminal domain catalyzes the reduction of the aldehyde to 3HP. The two domains can be produced independently and retain their enzymatic activity. This report focuses on the kinetic characterization of the C-terminal domain. Initial velocity patterns and inhibition studies showed the kinetic mechanism is ordered with NADPH binding first followed by malonyl-CoA. Malonic semialdehyde is released first, while CoA and NADP⁺ are released randomly. Analogs of malonyl-CoA showed that the thioester carbon is reduced, while the carboxyl group is needed for proper positioning. The enzyme transfers the pro-<em>S</em> hydrogen of NADPH to malonyl-CoA and pH rate profiles revealed that a residue with a pK_a value of about 8.8 must be protonated for activity. Kinetic isotope effects indicated that NADPH is not sticky (that is, NADPH dissociates from the enzyme faster than the rate of product formation) and product release is partially rate-limiting. Moreover, the mechanism is stepwise with the pH dependent step occurring before or after hydride transfer. The findings from this study will aid in the development of an eco-friendly biosynthesis of 3HP which is an industrial chemical used in the production of plastics and adhesives.</p></div>","PeriodicalId":8760,"journal":{"name":"Biochimica et biophysica acta. Proteins and proteomics","volume":"1872 5","pages":"Article 141033"},"PeriodicalIF":2.5,"publicationDate":"2024-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141632515","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}