Malate synthase from Streptomyces clavuligerus NRRL3585: cloning, molecular characterization and its control by acetate

Abstract

~~Malate synthase is a key enzyme of the glyoxylate cycle, which is an anaplerotic pathway essential for growth on acetate as the sole carbon source. The aceB gene, encoding malate synthase from Streptomyces clavuligems NRRL 3585, was cloned using PCR and fully sequenced. The ORF obtained encodes 541 amino acids with a deduced M, of 6OOOO, consistent with the' observed M, (62000-64000) of most malate synthase enzymes reported so far. The aceB gene has a high G+C content (71.5 molO/O), especially in the third codon position. A 50 bp region upstream of the malate synthase ORF was predicted to be a prokaryotic promoter region. The relationship between carbon source, antibiotic (cephalosporin) biosynthesis and malate synthase activity was investigated. Growth of S. clavuligerus on acetate as the major carbon source was delayed, compared to that on glycerol. Furthermore, high levels of malate synthase activity were associated with the presence of acetate in the growth medium. Growth on acetate also resulted in lower levels of cephalosporin production, compared to that on glycerol. The cloned 5. clavuligerus aceB gene was expressed in Escherichia coli BL2l (DE3). Transformants exhibited an approximately 713old increase in malate synthase activity, compared to the control, thereby demonstrating high-level expression of soluble and enzymically active malate synthase in the heterologous host.

Microbial community functional structure in response to antibiotics in pharmaceutical wastewater treatment systems

“…Fermentative antibiotic production wastewater commonly contains substrate residues (polysaccharide, cellulose, and hemicellulose), the target products (antibiotics), by-products like glyoxylate, malate and isocitrate and extraction solvents (Ayar-Kayali and Tarhan, 2006;Chan and Sim, 1998). Therefore, various carbon-degrading genes such as starch, cellulose/hemicellulose, chitin, lignin degradation genes, VanA, aceB, and amyA from the microbes in activated sludge are required to decompose these organic substances.…”

Mechanism of phosphate solubilization and antifungal activity of Streptomyces spp. isolated from wheat roots and rhizosphere and their application in improving plant growth

“…A 10-fold increase in MS activity and expression in Streptomyces mhcr0816 compared to the control (mhce0811) indicated the presence of MS isoforms that may have been active during the glyoxalate phase. Similar isoforms have been reported in cephalosporin-producing Streptomyces clavuligerus (Chan & Sim, 1998) and the excess malate thus produced could be secreted out of the cell probably via secondary transporter SAV1515 (Ikeda et al, 2003), which has a 1.5 kb nucleotide sequence [annotated in the whole genome sequence of Streptomyces avermitilis (Omura et al, 2001)]. SAV1515 is a secondary membrane transporter of the auxin efflux carrier (AEC) family assigned for IAA secretion.…”

Crotonyl-coenzyme A reductase provides methylmalonyl-CoA precursors for monensin biosynthesis by Streptomyces cinnamonensis in an oil-based extended fermentation

“…Analysis of the Streptomyces coelicolor genome sequence reveals the presence of genes putatively encoding isocitrate lyase and malate synthase, the two essential enzymes for this process. Malate synthase and isocitrate lyase have recently been studied from several streptomyces (Chan & Sim, 1998;Goh et al, 2003;Huttner et al, 1997;Loke & Sim, 2000;Loke et al, 2002;Soh et al, 2001). We have cloned and sequenced homologues of these genes from S. cinnamonensis.…”