Anti-peptide antibodies to the P4502D subfamily in rat, dog and man.
1. In order to obtain specific antibodies of the P4502D subfamily, we generated two anti-peptide antibodies against synthetic peptides, DPAQPPRD (peptide A) and DPTQPPRH (peptide B). The sequence of peptide A occurs in rat P4502D2, P4502D4 and human P4502D6, whereas the sequence of peptide B occurs in the dog P4502D subfamily. These sequences are closely related to an epitope of liver/kidney microsomal autoimmune hepatitis. 2. In immunoblotting studies, the anti-peptide antibody against peptide A recognized a 49-KDa protein in microsomes derived from human lymphoblasts expressing P4502D6 and rat liver. It showed no crossreactivity with microsomes from dog liver. In contrast, the anti-peptide antibody against peptide B recognized a 49-KDa protein only in microsomes of dog liver. These indicate that each anti-peptide antibody has the specificity for the respective sequences of the members of P4502D subfamily, with the species investigated herein. 3. In immunoinhibition studies, the anti-peptide antibodies against peptide B inhibited bunitrolol 4-hydroxylation and propranolol 4,5-hydroxylation, which are mediated by the dog P4502D subfamily. These data suggest that the anti-peptide antibodies against peptide B bind to the native and denatured forms of the P4502D subfamily. 4. The present study has demonstrated that the anti-peptide antibodies against this region are useful for studying the members of the P4502D subfamily.
Nakamura
A
A
Department of Toxicology, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Yamamoto
Y
Y
Tasaki
T
T
Sugimoto
C
C
Masuda
M
M
Kazusaka
A
A
Fujita
S
S
D000319:Adrenergic beta-Antagonists / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D015153:Blotting, Western; D019389:Cytochrome P-450 CYP2D6; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology* / Q000378:metabolism; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000276:immunology; D066298:In Vitro Techniques; D008214:Lymphocytes / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000276:immunology* / Q000378:metabolism; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology*; D011412:Propanolamines / Q000378:metabolism; D011433:Propranolol / Q000378:metabolism; D051381:Rats
D000319:Adrenergic beta-Antagonists; D000906:Antibodies; D007074:Immunoglobulin G; D010455:Peptides; D011412:Propanolamines; D003577:Cytochrome P-450 Enzyme System; D011433:Propranolol; D006899:Mixed Function Oxygenases; D019389:Cytochrome P-450 CYP2D6; C084811:bunitrolol