A mutant phospholipase D with enhanced thermostability from Streptomyces sp.
To investigate the contribution of amino acid residues to the thermostability of phospholipase D (PLD), a chimeric form of two Streptomyces PLDs (thermolabile K1PLD and thermostable TH-2PLD) was constructed. K/T/KPLD, in which residues 329-441 of K1PLD were recombined with the homologous region of TH-2PLD, showed a thermostability midway between those of K1PLD and TH-2PLD. By comparing the primary structures of Streptomyces PLDs, the seven candidates of thermostability-related amino acid residues of K1PLD were identified. The K1E346DPLD mutant, in which Glu346 of K1PLD was substituted with Asp by site-directed mutagenesis, exhibited enhanced thermostability, which was almost the same as that of TH-2PLD.
Hatanaka
Tadashi
T
Research Institute for Biological Sciences, Okayama, 7549-1 Kayo-cho, Jyobo-gun, Okayama 716-1241, Japan. hatanaka@bio-ribs.com
Negishi
Tomofumi
T
Mori
Koichi
K
D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D004795:Enzyme Stability; D006358:Hot Temperature; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010739:Phospholipase D / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
D011994:Recombinant Proteins; D010739:Phospholipase D