Structure-function studies of human aromatase.
Site-directed mutagenesis experiments have been carried out to determine the structure-function relationship of human aromatase. By sequence comparison, the region in aromatase that corresponds to the distal helix of cytochrome P-450cam has been identified to be Gln-298 to Val-313. Eight aromatase mutants with changes in this region, i.e. C299A, E302L, P308F, D309N, D309A, T310S, T310C, and S312C, have been generated using a mammalian cell stable-expression system. The results from site-directed mutagenesis studies indicate that the region containing Gln-298 to Val-313 is indeed a very important part of the active site of aromatase. The catalytic properties of P308F, D309N, and D309A have been examined in detail and are discussed. Active site-directed labeling is also an important approach to investigate the structure-function relationship of aromatase. HPLC-linked electrospray mass spectrometry is indicated as a useful technique for the characterization of active site-directed probe-modified enzyme. The mass spectral analysis of aromatase suggests that aromatase is glycosylated.
Chen
S
S
Division of Immunology, Beckman Research Institute of the City of Hope, Duarte, CA 91010.
Zhou
D
D
Swiderek
K M
KM
Kadohama
N
N
Osawa
Y
Y
Hall
P F
PF
D000595:Amino Acid Sequence; D000818:Animals; D001141:Aromatase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017433:Protein Structure, Secondary; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D001141:Aromatase