TY - JOUR
T1 - Purification and characterization of the hepatic CYP2C and 3A isozymes from phenobarbitone pretreated rhesus monkey
AU - Ramana, Kota V.
AU - Kohli, Krishan K.
N1 - Funding Information:
The authors thank Department of Atomic Energy, Bombay for financial support through grant 41/4/92-G, UGC, New Delhi for award of SRF to KVR, Rhone-Poulenc (India) Ltd, Bombay for the gift of sodium phenobarbitone and Dr. J.K. Batra of National Institute of Immunology, New Delhi for help in Western blotting and N-terminal sequencing.
PY - 1999
Y1 - 1999
N2 - Hepatic P450s, named M-3 and M-4 were purified from phenobarbitone pretreated rhesus monkey. These demonstrated polypeptide molecular mass of 50 and 52.5 kDa and specific content of 12 and 20 nmol P450/mg protein, respectively. Both the isozymes demonstrated low spin state of heme. Antibodies raised against M-3 inhibited the activity of aminopyrine, erythromycin and ethylmorphine N-demethylase in the microsomes obtained from PB pretreated rhesus monkey by 76, 40 and 35%, respectively. M-4 did the same by 69, 85 and 79%, respectively. These observations indicated M-3 and M-4 to be the members of CYP2C and 3A subfamilies, respectively. These results were substantiated by the observations that M-3 metabolized aminopyrine whereas M-4 metabolized aminopyrine, erythromycin and ethylmorphine in the reconstituted system. Microsomal lipids and cytochrome b5 enhanced the rate of these reactions. Further confirmation to the identity of these isozymes was provided by N-terminal amino acid sequences. The first 10 N-terminal amino acid residues of M-3 were 90% similar to CYP2C20 and 2C9 and that of M-4 were 100 and 90% similar to CYP3A8 and 3A5, respectively. In conclusion, two isozymes of hepatic P450 purified from PB pretreated rhesus monkey belong to CYP2C and 3A subfamilies.
AB - Hepatic P450s, named M-3 and M-4 were purified from phenobarbitone pretreated rhesus monkey. These demonstrated polypeptide molecular mass of 50 and 52.5 kDa and specific content of 12 and 20 nmol P450/mg protein, respectively. Both the isozymes demonstrated low spin state of heme. Antibodies raised against M-3 inhibited the activity of aminopyrine, erythromycin and ethylmorphine N-demethylase in the microsomes obtained from PB pretreated rhesus monkey by 76, 40 and 35%, respectively. M-4 did the same by 69, 85 and 79%, respectively. These observations indicated M-3 and M-4 to be the members of CYP2C and 3A subfamilies, respectively. These results were substantiated by the observations that M-3 metabolized aminopyrine whereas M-4 metabolized aminopyrine, erythromycin and ethylmorphine in the reconstituted system. Microsomal lipids and cytochrome b5 enhanced the rate of these reactions. Further confirmation to the identity of these isozymes was provided by N-terminal amino acid sequences. The first 10 N-terminal amino acid residues of M-3 were 90% similar to CYP2C20 and 2C9 and that of M-4 were 100 and 90% similar to CYP3A8 and 3A5, respectively. In conclusion, two isozymes of hepatic P450 purified from PB pretreated rhesus monkey belong to CYP2C and 3A subfamilies.
KW - CYP2C
KW - CYP3A
KW - Cytochrome P450
KW - Phenobarbitone
KW - Rhesus monkey
UR - http://www.scopus.com/inward/record.url?scp=0032849211&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0032849211&partnerID=8YFLogxK
U2 - 10.1023/A:1006902212598
DO - 10.1023/A:1006902212598
M3 - Article
C2 - 10497881
AN - SCOPUS:0032849211
SN - 0300-8177
VL - 198
SP - 79
EP - 88
JO - Molecular and Cellular Biochemistry
JF - Molecular and Cellular Biochemistry
IS - 1-2
ER -