Purification and characterization of [3H]mepyramine (histamine H1 antagonist)-binding protein from rat liver: a highly homologous protein with cytochrome P450 2D.
A protein having a high-affinity binding site for [3H]mepyramine (MBP) was purified to homogeneity from rat liver membranes. The purified MBP has a single type of binding site for [3H]mepyramine with Kd value of 18.5 nM, and its molecular weight was determined to be 56,000 by SDS polyacrylamide gel electrophoresis. Amino acid sequences of twelve tryptic peptides derived from MBP are highly homologous with those of rat debrisoquine 4-hydroxylase (cytochrome P450 2D1) and other rat P450 2D subfamily members. In immunoblotting analysis, an antibody against rat P450 2D1 stained a band corresponding to MBP with Mr of 56,000; its migration position was clearly different from that of rat P450 2D1. Substrates and inhibitors of debrisoquine 4-hydroxylase potently displace [3H]-mepyramine binding to MBP. Quinine and quinidine showed 400 and 80 times, respectively, higher affinity for MBP than for debrisoquine 4-hydroxylase. These results suggest that MBP is a novel P450 2D family member.
Fukui
H
H
Department of Pharmacology II, Faculty of Medicine, Osaka University.
Mizuguchi
H
H
Liu
Y Q
YQ
Wang
N P
NP
Hayashi
H
H
Kangawa
K
K
Wakamiya
T
T
Leurs
R
R
Shiba
T
T
Matsuo
H
H
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism; D006634:Histamine H1 Antagonists / Q000737:chemistry* / Q000302:isolation & purification*; D015151:Immunoblotting; D008099:Liver / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011738:Pyrilamine / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014316:Tritium
D002352:Carrier Proteins; D065607:Cytochrome P-450 Enzyme Inhibitors; D006634:Histamine H1 Antagonists; D008565:Membrane Proteins; D010446:Peptide Fragments; D014316:Tritium; D003577:Cytochrome P-450 Enzyme System; D011738:Pyrilamine