Objectives: Alteration of the cofactor specificity of acrylyl-CoA reductase (AcuI) catalyzing the NAD(P)H-dependent reduction of acrylyl-CoA to propionyl-CoA is often desirable for designing of artificial metabolic pathways of various appointments.
Results: Several variants of AcuIs from Escherichia coli K-12 with multiple amino acid substitutions to alter the cofactor preference were obtained by site directed mutagenesis and the modified enzymes as His-tagged proteins were characterized. The simultaneous substitutions of arginine-180, arginine-198 and serine-178 residues by alanine in the enzyme pocket sequence as well as other amino acid changes decreased both NADPH- and NADH-dependent activities in comparison to the wild-type enzyme. The replacement of serine-156 by glutamic acid decreased NADPH-dependent activity at least 7000-fold but NADH-dependent activity only by threefold. The replacement of serine-156 by aspartic acid decreased NADPH-dependent activity 70-fold with fair preservation of activity and specificity to NADH.
Conclusions: These results demonstrated a relevance of Asp156 in the interaction of AcuI from E. coli K-12 with NADH as a coenzyme. These findings may provide reference information for shifting coenzyme specificity of acrylyl-CoA reductases.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1007/s10529-021-03130-0 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Dioxin-elicited decrease in cobalamin redirects propionyl-CoA metabolism to the β-oxidation-like pathway resulting in acrylyl-CoA conjugate buildup.
J Biol Chem
September 2022
Biochemistry & Molecular Biology, Michigan State University, East Lansing, Michigan, USA; Institute for Integrative Toxicology, Michigan State University, East Lansing, Michigan, USA. Electronic address:
2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a persistent environmental contaminant that induces diverse biological and toxic effects, including reprogramming intermediate metabolism, mediated by the aryl hydrocarbon receptor. However, the specific reprogramming effects of TCDD are unclear. Here, we performed targeted LC-MS analysis of hepatic extracts from mice gavaged with TCDD.
View Article and Find Full Text PDFAlteration of cofactor specificity of the acrylyl-CoA reductase from Escherichia coli.
Biotechnol Lett
July 2021
Laboratory of Methylotrophy, Federal Research Center, "Pushchino Scientific Center for Biological Research, Russian Academy of Sciences" IBPM RAS, Pushchino, Moscow Region, Russia, 142290.
Objectives: Alteration of the cofactor specificity of acrylyl-CoA reductase (AcuI) catalyzing the NAD(P)H-dependent reduction of acrylyl-CoA to propionyl-CoA is often desirable for designing of artificial metabolic pathways of various appointments.
Results: Several variants of AcuIs from Escherichia coli K-12 with multiple amino acid substitutions to alter the cofactor preference were obtained by site directed mutagenesis and the modified enzymes as His-tagged proteins were characterized. The simultaneous substitutions of arginine-180, arginine-198 and serine-178 residues by alanine in the enzyme pocket sequence as well as other amino acid changes decreased both NADPH- and NADH-dependent activities in comparison to the wild-type enzyme.
β-alanine biosynthesis in Methanocaldococcus jannaschii.
J Bacteriol
August 2014
Department of Biochemistry, Virginia Polytechnic Institute and State University, Blacksburg, Virginia, USA
One efficient approach to assigning function to unannotated genes is to establish the enzymes that are missing in known biosynthetic pathways. One group of such pathways is those involved in coenzyme biosynthesis. In the case of the methanogenic archaeon Methanocaldococcus jannaschii as well as most methanogens, none of the expected enzymes for the biosynthesis of the β-alanine and pantoic acid moieties required for coenzyme A are annotated.
View Article and Find Full Text PDFAcrylyl-coenzyme A reductase, an enzyme involved in the assimilation of 3-hydroxypropionate by Rhodobacter sphaeroides.
J Bacteriol
October 2013
Department of Microbiology, The Ohio State University, Columbus, Ohio, USA.
The anoxygenic phototroph Rhodobacter sphaeroides uses 3-hydroxypropionate as a sole carbon source for growth. Previously, we showed that the gene (RSP_1434) known as acuI, which encodes a protein of the medium-chain dehydrogenase/reductase (MDR) superfamily, was involved in 3-hydroxypropionate assimilation via the reductive conversion to propionyl-coenzyme A (CoA). Based on these results, we speculated that acuI encoded acrylyl-CoA reductase.
View Article and Find Full Text PDFStudies of human mitochondrial 2,4-dienoyl-CoA reductase.
Arch Biochem Biophys
February 2005
Department of Biology and Chemistry, City University of Hong Kong, 83 Tat Chee Avenue, Kowloon, Hong Kong SAR, PR China.
Mitochondrial 2,4-dienoyl-CoA reductase is a key enzyme for the beta-oxidation of unsaturated fatty acids. Sequence alignment indicates that there are five highly conserved acidic residues, one of which might act as a proton donor. We constructed five mutant expression plasmids of human mitochondrial 2,4-dienoyl-CoA reductase using site-directed mutagenesis.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!