title: Enhanced stability of newly isolated trimeric l-methionine-N-carbamoylase from Brevibacillus reuszeri HSN1 by covalent immobilization.
abstract: Newly isolated and partially purified trimeric l-methionine-N-carbamoylase from Brevibacillus reuszeri HSN1 was immobilized by covalent coupling to a well-known support material, Eupergit® C. Approximately 80% enzyme activity yield was achieved with ≈61% binding of a soluble protein from a solution containing 5 mg/mL protein. The immobilized preparation was found to be quite unstable due to a poor multisubunit covalent interaction of trimeric enzyme. Additional cross-linking with polyaldehyde-dextran was done to sustain the biotechnological application of immobilized enzyme. The temperature and pH optima of immobilized enzyme were increased by 10°C and 0.5 unit, respectively. The enzyme was significantly stabilized and retained ≈93% enzyme activity when incubated at 60°C for 60 Min, whereas free enzyme lost ≈50% activity. It was recycled nine times with ≈100% conversion efficiency when batch experiments were carried out at 35°C, pH 7.5, for the 180 Min cycle, using 5% N-carbamoyl-l-methionine as the substrate. The half-life of the immobilized preparation was determined as 23 cycles and is significant. Approximately 50% of enzyme activity was retained even after 5 months of storage at 4°C, whereas free enzyme lost complete enzyme activity. Hence, we could enhance the stability of l-methionine-N-carbamoylase to make it a potential biocatalyst for biotechnological production of α-amino acids.
authors.affiliation: CSIR-Institute of Microbial Technology, Sector 39-A, Chandigarh, India. hemraj@imtech.res.in
authors.forename: Hemraj S
authors.initials: HS
authors.lastname: Nandanwar
authors.forename: Rakesh M
authors.initials: RM
authors.lastname: Vohra
authors.forename: Gurinder S
authors.initials: GS
authors.lastname: Hoondal
mesh_terms: D058675:Brevibacillus / Q000378:metabolism*; D002219:Carbamates / Q000737:chemistry* / Q000378:metabolism*; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism*; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008715:Methionine / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000378:metabolism*; D011108:Polymers / Q000737:chemistry; D013696:Temperature
chemical_list: D002219:Carbamates; D004800:Enzymes, Immobilized; D011108:Polymers; C072457:N-carbamylmethionine; C065843:Eupergit C; D008715:Methionine