The CYP152 family of cytochrome P450 enzymes (P450s or CYPs) are bacterial peroxygenases that use hydrogen peroxide to drive hydroxylation and decarboxylation of fatty acid substrates. We have expressed and purified a novel CYP152 family member – CYP152K6 from the methylotroph Bacillus methanolicus MGA3. CYP152K6 was characterized using spectroscopic, analytical and structural methods. CYP152K6, like its peroxygenase counterpart P450_SPα (CYP152B1) from Sphingomonas paucimobilis, does not undergo significant fatty acid-induced perturbation to the heme spectrum, with the exception of a minor Soret shift observed on binding dodecanoic acid. However, CYP152K6 purified from an E. coli expression system was crystallized and its structure was determined to 1.3 Å with tetradecanoic acid bound. No lipids were present in conditions used for crystallogenesis, and thus CYP152K6 must form a complex by incorporating the fatty acid from E. coli cells. Turnover studies with dodecanoic acid revealed several products, with 2-hydroxydodecanoic acid as the major product and much smaller quantities of 3-hydroxydodecanoic acid. Secondary turnover products were undec-1-en-1-ol, 2-hydroxydodec-2-enoic acid and 2,3-dihydroxydodecanoic acid. This is the first report of a 2,3-hydroxylated fatty acid product made by a peroxygenase P450, with the dihydroxylated product formed by CYP152K6-catalyzed 3-hydroxylation of 2-hydroxydodecanoic acid, but not by 2-hydroxylation of 3-hydroxydodecanoic acid.

CYP152 细胞色素 P450 酶家族（P450 或 CYP）是细菌过氧化酶，它使用过氧化氢来驱动脂肪酸底物的羟基化和脱羧。我们从甲基营养菌甲醇芽孢杆菌MGA3 中表达并纯化了一个新的 CYP152 家族成员——CYP152K6。CYP152K6 使用光谱、分析和结构方法进行表征。CYP152K6 与其来自Sphingomonas paucimobilis 的过氧合酶对应物 P450 _SPα (CYP152B1)一样，除了在结合十二烷酸时观察到轻微的 Soret 位移外，不会对血红素谱造成显着的脂肪酸诱导扰动。然而，从大肠杆菌中纯化的 CYP152K6表达系统结晶，其结构被确定为 1.3 Å，结合了十四烷酸。在用于结晶发生的条件下不存在脂质，因此 CYP152K6 必须通过结合来自大肠杆菌细胞的脂肪酸形成复合物。十二烷酸的营业额研究揭示了几种产品，其中 2-羟基十二烷酸是主要产品，而 3-羟基十二烷酸的数量要少得多。次生周转产品为 undec-1-en-1-ol、2-hydroxydodec-2-enoic acid 和 2,3-dihydroxydodecanoic acid。这是由过氧合酶 P450 制备的 2,3-羟基化脂肪酸产物的首次报道，其中二羟基化产物由 CYP152K6 催化的 2-羟基十二烷酸的 3-羟基化而不是由 3-羟基十二烷酸的 2-羟基化形成.