Molecular Cloning and Characterization of the Gene Encoding Cold-Active β-Galactosidase from a Psychrotrophic and Halotolerant Planococcus sp. L4

• DOI: 10.1021/jf062910r
• Publication Date: 2007-02-28T12:21:58Z
• Authors: Ji M. Hu; He Li; Li X. Cao; Pei C. Wu; Chen T. Zhang; Shu L. Sang; Xiao Y. Zhang; Min J. Chen; Jia Q. Lu; Yu H. Liu
• Keywords: Molecular mass; Halotolerance; Thermolabile

The gene bgaP encoding cold-active beta-galactosidase from a psychrotrophic and halotolerant Planococcus sp. L4 was cloned into Escherichia coli sequenced. A sequence analysis of the DNA responsible for BgaP revealed an open reading frame 2031 bp protein 677 amino acid residues. heterologously expressed in E. purified followed by Ni2+ affinity chromatography. molecular mass native enzyme approximately 156 kDa as determined gel filtration. results sodium dodecyl sulfate-polyacrylamide electrophoresis deduced indicated masses 78 77.311 kDa, respectively, suggesting that is dimer. had isoelectric point 4.8 exhibited maximal activity at 20 degrees C pH 6.8 under assay conditions used. particularly thermolabile, losing all only 10 min 45 C. It able to hydrolyze lactose substrate, well o-nitrophenyl-beta-D-galactopyranoside (ONPG); Km values with ONPG were calculated be 5.4 20.4 mM 5 C, respectively. catalytic efficiencies BagP 14 47 times more than Therefore, could conceivably developed fulfill practical requirements enable its use removal milk dairy products low temperature or reporter psychrophilic genetic systems.