Human cytomegalovirus expresses an unusual protein kinase UL97, a member of ${H_V}{U_L}$ family of protein kinase. UL97 can phosphorylate nucleoside analogs such as ganciclovir as well as protein/peptide. It has previously been reported that UL97 is able to phosphorylate a KESYSVYVYKV peptide and that P+5 position (K) is important. We examined the extent of contribution of other positions (P-4 through P+6) of the peptide to be substrate of UL97 using alanine substituted peptides (Ala scanning) and deleted peptides. The result suggested that the E (P-2) is negative effect and P+5 (K) is still important. The peptide YSVYVYK is the shortest substrate enough to show high activity, which could be a starting point to develop peptidomimetic drug. This study would give important information to deeply understand the substrate specificity of UL97 and develop an antiviral drug using the small peptide identified here.