Characteristics of DNA-binding activity of human cytomegalovirus ppUL44

Human cytomegalovirus (HCMV)-specific monoclonal antibody, SCMVM34, recogni zes the early antigen encoded by UL44 of HCMV, This antigen is confined to the nucleus of HCMV-infected cells. This study was performed to characteriz e the DNA-binding activity of the protein encoded by UL44 of HCMV, The nucl ear and cytoskeletal fraction of HCMV-infected cells was subjected to 0.4 M NaCl extraction, DEAE-Sephacel ion exchange chromatography, DNA-cellulose chromatography and SDS-PAGE analysis with monitoring of the reactive protei n using SCMVM34 monoclonal antibody. The molecular weights of the resultant proteins were found to be 34, 40 and 52 kDa. The internal peptide fragment s were isolated by tryptic digestion and reverse-phase HPLC, The internal a mino acid sequence analysis of the peptides from the HPLC profile revealed that the antigen recognized by SCMVM34 monoclonal antibody was ppUL44, The reactive antigen began to be eluted from 250 mM NaCl (Tris-HCl pH 7.4) in D NA cellulose. The 34 kDa protein seems to bind to DEAE more tightly than th e 52 kDa protein. The surface charge of 34 kDa might be more basic, Conclus ively, the antigen recognized by SCMVM34 was the protein encoded by HCMV UL 44, which was localized in the nuclei after HCMV infection, and was the DNA -binding protein with the characteristic that the surface charge of the mol ecule was more basic, as the molecular weights of the protein were decrease d.