Structure of an activity suppressing Fab fragment to cytochrome P450 aromatase: insights into the antibody-antigen interactions.
The crystal structure of a Fab fragment (Fab3-2C2) of a monoclonal antibody raised against aromatase cytochrome P450 P450arom) has been determined at 3.0 A resolution. P450arom is a membrane bound enzyme responsible for the catalysis of indrogens to estrogens, the process of aromatization, and hence has been implicated in hormone-dependent breast cancer. The Fab fragment of MAb3-2C2 IgG suppresses P450arom activity in a dose dependent manner. The Fab3-2C2 molecule crystallizes n the space group P2(1)2(1)2(1) with a unit cell of a= 154.89 A, b = 73.51 A, and c= 36.90 A. The crystal structure consists of a light and a heavy chain in the asymmetric unit, each characterized by the greek-key antiparallel beta barrel folding seen in all Fab structures. The average elbow angle between the two domains is 143 degrees. Modeling of the interactions between the variable domains of the antibody and a known model of P450arom maps the epitope to a region of the enzyme that is consistent with the available biochemical data and the activity-suppressing function of the antibody. The epitope mapping result is further supported by the inability of MAb3-2C2 IgG to suppress the activity of, or to interact with placental porcine P450arom, which is 81% identical (86% similar) to human P450arom but has a few key substitutions in the putative epitope region.
Sawicki
M W
MW
Roswell Park Cancer Institute, Buffalo, NY 14263, USA.
Ng
P C
PC
Burkhart
B M
BM
Pletnev
V Z
VZ
Higashiyama
T
T
Osawa
Y
Y
Ghosh
D
D
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D000937:Antigen-Antibody Reactions; D001141:Aromatase / Q000276:immunology* / Q000302:isolation & purification; D047072:Aromatase Inhibitors; D001483:Base Sequence; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification; D007074:Immunoglobulin G / Q000737:chemistry / Q000276:immunology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D013552:Swine
D000911:Antibodies, Monoclonal; D047072:Aromatase Inhibitors; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D001141:Aromatase