THE MOLECULAR STRUCTURE OF HUMAN CYTOMEGALOVIRUS ASSEMBLIN - A NOVEL SERINE PROTEASE AND AN UNIQUE DRUG TARGET Huey-Sheng Shieh, William C. Stallings, Ravi G. Kurumbail, Anna M. Stevens, Jina Y. Pak, Eric J. Sturman, Roderick A. Stegeman, Mark O. Palmier, Arthur J. Wittwer, and Barry C. Holwerda, Searle/Monsanto, BB4K, 700 Chesterfield Parkway N., Chesterfield, MO 63198, USA

Human cytomegalovirus (HCMV), a herpesvirus, is a ubiquitous opportunistic pathogen which causes clinically apparent disease in congenitally infected infants and in immunocompromised and immunosuppressed adults. All members of the herpesvirus family encode a unique serine protease, assemblin, which is necessary for viral replication. HCMV assemblin was crystallized in space group P4₂ with a dimer in each asymmetric unit. The crystals diffract to 2.2 Å resolution. The structure was determined at 2.8 Å resolution by multiple isomorphous replacement methods combined with solvent flattening and averaging. The assemblin molecule folds in a eight-stranded barrel which is stabilized by surrounding [[alpha]] helices. One of the exterior surfaces of the barrel contains the constellation of residues that forms the active site. This folding is quite different from the other classes of serine protease, such as the two-domain structures of chymotrypsin series, and the one-domain structures of subtilisin and serine carboxypeptidase II series. The primary feature of the dimer interface in the assemblin is the [[alpha]] helix (residues 217 235) that runs parallel to the dimer dyad and is recognized by a groove formed by three helices from the other subunit of the dimer. The residues around the putative internal cleavage sites (143 and 209) consistently show poor electron density in various Fourier maps and have not been modelled in the current structure. The structure has been used to identify residues that may be involved in catalysis and the role of these residues has been investigated by site directed mutagenesis. These results will also be presented.