THREE DIMENSIONAL STRUCTURE OF HUMAN ORNITHINE AMINOTRANSFERASE: CHARGE DISTRIBUTION AT THE ACTIVE SITE EXPLAINS THE SELECTIVITY FOR W - GROUP OF DIBASIC AMINO ACIDS. Betty W. Shen^*, Michael Hennig^#, Erhard Hohenester^#, and Johan N. Jansonius^#.^*Department of Molecular of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT USA; ^#Biozentrum, University of Basel, Basel CH4056 Switzerland

Ornithine aminotransferase (OAT) catalyses exclusively thetransfer of the delta-amino group of ornithine to alpha-ketoglutarate, forming glutamate-r-semialdyhyde and glutamic acid. It is one of the major matabolic enzymes that modulate nitrogen fixation in the urea cycle as well as the synthesis of argine, proline, and spermidine.

Inhibition of OAT has recently been explored as means of detoxification in animal experiments. In view of its diversified roles, it is of interest to compare the structure of OAT with other subgroups of the transaminase superfamily in order to understand the basis of its extraordinary specificity.

The structure of OAT is presented at 2.5Å resolution. The recombinant enzyme crystallized in the trigonal space group P3(2)21 with one and a half dimers per asymmetric unit. The refined model has a R-factor of 17.9% (free-R = 23.3%) and excellent stereochemistry. Each subunit of the dimeric enzyme contains three distinct regions: an extended N-terminal segment, a cofactor binding large domain, and a relatively flexible small domain. The OAT dimer is stabilized by a large contact surface between the large domains of the two subunits and between the N-terminal segment of one sununit with the large domian of the other. The pyridoxal-5'-phosphate (PLP) cofactor is bound to the large domain close to the subunit interface and is stabilized through multiple hydrophobic and electrostatic interactions to both subunits. Glu235 and Arg413, which form a salt bridge in at the entrance to the active site pocket, are the most probable residues for the binding of ornithine. The location of Glu235 explains the selectivity of OAT for the delta-amino group despite the presence of the more reactive alpha-amino groups of ornithine and glutamate.