cDNA and deduced amino acid sequences of a dog hepatic cytochrome P450IIB responsible for the metabolism of 2,2',4,4',5,5'-hexachlorobiphenyl.
cDNA and deduced amino acid sequences of a dog hepatic cytochrome P450IIB responsible for the metabolism of 2,2',4,4',5,5'-hexachlorobiphenyl.
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负责 2,2,4,4,5,5-六氯联苯代谢的狗肝细胞色素 P450IIB 的 cDNA 和推导的氨基酸序列。
DOI：
10.1016/0003-9861(90)90419-y
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发表时间：
1990
影响因子：
3.9
通讯作者：
Halpert,JR
中科院分区：
文献类型：
--
作者：
Graves,PE;Elhag,GA;Ciaccio,PJ;Bourque,DP;Halpert,JR
测定了苯巴比妥（PB）诱导的雄性比格犬肝细胞色素P450的cDNA序列。利用兔P450IIB cDNA探针(R. Gasser， M. Negishi, and R. M. Philpot, 1988；从pb处理犬的肝脏mRNA制备的λgt11文库中分离到一个2.6千碱基对插入的cDNA克隆。对克隆的插入片段进行测序，发现包含一个开放阅读框，编码494个氨基酸的多肽（Mr56,183）。根据与其他物种的细胞色素P450的同源性，该编码蛋白可以被分配到P450IIB亚家族。推导出的氨基酸序列与兔P450 B0 （P450IIB4）的同源性为79%，与大鼠P450b （P450IIB1）的同源性为75%。与其他P450II亚家族相比，狗的序列一致性下降到52%以下。狗cDNA编码的nh2末端30个氨基酸与纯化犬细胞色素P450 PBD-2的序列分析相同，氨基酸组成与PBD-2蛋白的氨基酸组成一致(D. B. Duignan， I. G. Sipes， T. B. Leonard, and J. R. Halpert, 1987,Arch。物化学。Biophys.255, 290 - 303)。Northern blots检测到两个长度分别为1.9和2.9千碱基的mRNA物种，它们与狗P450IIB cDNA编码区杂交。与未处理的动物相比，pb处理的狗肝脏中总可杂交mRNA水平增加了约6倍。这种增加与报道的PBD-2蛋白水平增加近6倍和2,2 ',4,4 ',5,5 ' -六氯联苯治疗后肝脏代谢率增加5倍密切相关。这两种mRNA可能是由位于3 ' -非编码区不同的聚腺苷化信号或多个PBD-2基因的转录引起的。Southern blot分析表明，狗P450IIB亚家族包含至少两个密切相关的基因。
The nucleotide sequence of a cDNA that codes for the major phenobarbital (PB)-inducible male beagle dog hepatic cytochrome P450 has been determined. Using a rabbit P450IIB cDNA probe (R. Gasser, M. Negishi, and R. M. Philpot, 1988,Mol. Pharmacol.32, 22–30), a cDNA clone with a 2.6-kilobase pair insert was isolated from a λgt11 library prepared from hepatic mRNA from a PB-treated dog. The cloned insert was sequenced and found to contain an open reading frame coding for a polypeptide of 494 amino acids (Mr56,183). The encoded protein can be assigned to the P450IIB subfamily on the basis of homology to cytochromes P450 from other species. The deduced amino acid sequence is 79% identical to that reported for rabbit P450 B0 (P450IIB4) and 75% identical to that for rat P450b (P450IIB1). The sequence identity decreases to less than 52% when the dog sequence is compared with other P450II subfamilies. The deduced NH2-terminal 30 amino acids encoded by the dog cDNA are identical to those determined by sequence analysis of purified dog cytochrome P450 PBD-2, and the amino acid composition concurs with that determined for the PBD-2 protein (D. B. Duignan, I. G. Sipes, T. B. Leonard, and J. R. Halpert, 1987,Arch. Biochem. Biophys.255, 290–303). Northern blots revealed two mRNA species of approximately 1.9 and 2.9 kilobases in length, which hybridized to the coding region of the dog P450IIB cDNA. The level of total hybridizable mRNA was increased approximately sixfold in livers from PB-treated dogs compared with that in untreated animals. This increase correlates well with the reported nearly sixfold increase in the level of PBD-2 protein and the fivefold increase in the rate of hepatic metabolism of 2,2′,4,4′,5,5′-hexachlorobiphenyl following PB treatment. The two mRNA species may result from the use of different polyadenylation signals located in the 3′-noncoding region or from transcription of more than one gene for PBD-2. Southern blot analysis indicated that the dog P450IIB subfamily contains at least two closely related genes.