Imaoka S, Funae Y
Laboratory of Chemistry, Osaka City University Medical School.
J Biochem. 1990 Jul;108(1):33-6. doi: 10.1093/oxfordjournals.jbchem.a123157.
The pulmonary cytochrome P-450, P450 L-2, was purified 460-fold from pulmonary microsomes of untreated male rats. Its specific content was 10.6 nmol/mg of protein. The monomeric molecular weight was 54,000 on SDS-polyacrylamide gel electrophoresis. The CO-reduced absorption maximum of P450 L-2 was at 451 nm, and the oxidized heme iron appeared to be in the low-spin state, as deduced from the Soret maximum at 421 nm. P450 L-2 had high lauric acid omega- and (omega-1)-hydroxylation activities, but low prostaglandin A1 omega- and (omega-1)-hydroxylation activities. It catalyzed the O-dealkylation of 7-ethoxycoumarin, but was not efficient in the hydroxylation of testosterone or the N-demethylation of aminopyrine. The NH2-terminal amino acid sequence of P450 L-2 was V-L-N-F-L-X-P-X-L (X being an unidentified residue). The catalytic properties of P450 L-2 resembled those of P450 K-5, the major rat renal cytochrome P-450. However, anti-P450 K-5 antibody did not cross-react with P450 L-2, and these forms had different NH2-terminal sequences. To judge from the results of NH2-terminal sequence analysis, P450 L-2 seems to be placed in the IVB gene family. Also, P-450 IIB1 was detected by immunoblotting in one of the peaks on ion-exchange HPLC during the purification of P450 L-2, suggesting the presence of P-450 IIB1 in rat pulmonary microsomes.
从未经处理的雄性大鼠肺微粒体中纯化出肺细胞色素P-450（P450 L-2），纯化倍数为460倍。其比含量为10.6 nmol/mg蛋白质。在SDS-聚丙烯酰胺凝胶电泳上，单体分子量为54,000。P450 L-2的一氧化碳还原吸收最大值在451 nm处，从421 nm处的Soret最大值推断，氧化血红素铁似乎处于低自旋状态。P450 L-2具有较高的月桂酸ω-和（ω-1）-羟基化活性，但前列腺素A1的ω-和（ω-1）-羟基化活性较低。它催化7-乙氧基香豆素的O-脱烷基反应，但对睾酮的羟基化或氨基比林的N-脱甲基反应效率不高。P450 L-2的NH2-末端氨基酸序列为V-L-N-F-L-X-P-X-L（X为未鉴定的残基）。P450 L-2的催化特性与大鼠主要肾细胞色素P-450即P450 K-5的催化特性相似。然而，抗P450 K-5抗体与P450 L-2不发生交叉反应，且这些形式具有不同的NH2-末端序列。从NH2-末端序列分析结果判断，P450 L-2似乎属于IVB基因家族。此外，在纯化P450 L-2的离子交换高效液相色谱的一个峰中，通过免疫印迹检测到了P-450 IIB1，这表明大鼠肺微粒体中存在P-450 IIB1。
