Kinetic and immunochemical studies of a receptor-like protein that binds aromatic hydrocarbons.
A 29-kDa cytosolic protein that binds polycyclic aromatic hydrocarbons (PBP) with high affinity, specificity and saturability was identified in and was purified from C57BL/6J mouse liver (Collins, S., and Marletta, M. A. (1986) Biochemistry 25, 4322-4329). Kinetic studies showed that benzo[a]pyrene had a koff = 0.28 +/- 0.06 min-1 and a kon = 7.0 +/- 0.2 x 10(7) M-1 min-1. The Kd calculated from these rates is 4.0 +/- 0.9 nM which agrees with equilibrium measurements. This provides support for the description of this ligand-protein interaction as simple mass action binding of one ligand per protein molecule. Polyclonal antiserum with a high titer of antibodies specific for this protein was prepared. Western blot analyses from mouse tissues showed highest levels in liver, heart, kidney, and lung with lower levels in intestine, spleen, thymus, testes, and brain. Serum was negative. Liver cytosols from 7-week-old males of eight strains of mice (C57Bl/6J, BALB/cJ, C3H/HeJ, A/J, CD1/Cr, DBA/2J, AKR/J, and SWR/J) had similar concentrations of the PBP. The PBP was also present in liver cytosol from 7-week-old female and 42-week-old male C57Bl/6J mice. High affinity specific binding for benzo[a]pyrene was also found in rat, rabbit, guinea pig, scup, and chicken liver cytosols. This binding had low nM equilibrium binding constants and was not competed by tetrachlorodibenzofuran distinguishing these sites from the Ah receptor. These studies show that the PBP or similar receptor-like proteins are widespread among mouse tissues and strains and evolutionarily diverse animal species.
Barton
H A
HA
Department of Applied Biological Sciences, Massachusetts Institute of Technology, Cambridge 02139.
Marletta
M A
MA
D000818:Animals; D000918:Antibody Specificity; D001564:Benzo(a)pyrene / Q000378:metabolism; D001580:Benzopyrenes / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D002645:Chickens; D003600:Cytosol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005399:Fishes; D006168:Guinea Pigs; D007106:Immune Sera / Q000276:immunology; D007118:Immunoassay; D008099:Liver / Q000032:analysis*; D008297:Male; D051379:Mice; D008805:Mice, Inbred A; D008806:Mice, Inbred AKR; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D011817:Rabbits; D051381:Rats; D013045:Species Specificity; D014018:Tissue Distribution
D001580:Benzopyrenes; D007106:Immune Sera; D001564:Benzo(a)pyrene