Hayashi Asuka, Saitou Hiroki, Mori Tomomi, Matano Ikue, Sugisaki Hiroyuki, Maruyama Kiyofumi
Department of Biomolecular Science, Faculty of Engineering, Gifu University, Gifu, Japan.
Biosci Biotechnol Biochem. 2012;76(3):559-66. doi: 10.1271/bbb.110860.
Monoacetylphloroglucinol (MAPG) acetyltransferase, catalyzing the conversion of MAPG to 2,4-diacetylphloroglucinol (DAPG), was purified from Pseudomonas sp. YGJ3 grown without Cl(-). Cl(-) and pyoluteorin repressed expression of the enzyme. SDS-polyacrylamide gel electrophoresis showed that the purified enzyme (M(r)=330 kDa) was composed of three subunits of 17, 38, and 43 kDa, and protein sequencing identified these as PhlB, PhlA, and PhlC respectively. The enzyme catalyzed the reversible disproportionation of 2 moles of MAPG to phloroglucinol (PG) and DAPG. The equilibrium constant K (=[DAPG][PG]/MAPG) was estimated to be about 1.0 at 25 °C. A KpnI 20-kb DNA fragment was cloned from the genomic DNA of strain YGJ3, and a 12,598-bp long DNA region containing the phl gene cluster phlACBDEFGHI was sequenced. PCR cloning and expression of the phl genes in Escherichia coli confirmed that expression of phlACB genes produced MAPG ATase.
单乙酰间苯三酚（MAPG）乙酰转移酶可催化MAPG转化为2,4 - 二乙酰间苯三酚（DAPG），该酶是从在无Cl(-)条件下生长的假单胞菌属YGJ3中纯化得到的。Cl(-)和绿脓菌素可抑制该酶的表达。SDS - 聚丙烯酰胺凝胶电泳显示，纯化后的酶（M(r)=330 kDa）由17 kDa、38 kDa和43 kDa的三个亚基组成，蛋白质测序分别将它们鉴定为PhlB、PhlA和PhlC。该酶催化2摩尔MAPG可逆歧化为间苯三酚（PG）和DAPG。在25℃时，平衡常数K（=[DAPG][PG]/MAPG）估计约为1.0。从菌株YGJ3的基因组DNA中克隆出一个KpnI 20 - kb的DNA片段，并对包含phl基因簇phlACBDEFGHI的12,598 - bp长的DNA区域进行了测序。phl基因在大肠杆菌中的PCR克隆和表达证实，phlACB基因的表达产生了MAPG ATase。
