The Phosphorylation is the key activity of nm23-H1. The aim of this study is to explore the effect of different amino acid mutation on the phosphorylation status of nm23-H1.The wild type nm23-H1 was as the control of this study. Autoradiography was used for detecting the serine and histidine autophosphorylation of wild type (WT) and mutant nm23-H1 (P96S, H118F, S120G and S44A); RP-HPLC was used for detecting the NDPK activity of above proteins.The autophosphorylation activities of serine and histidine from high to low were P96S, WT, S44A, S120G and H118F, respectively, while the NDPK activities from high to low were WT, S120G, P96S, S44A, H118F. A highly positive correlation was found between serine and histidine autophosphorylation activity of above proteins (r =0.985, P 0.05, and r =0.482, P >0.05, respectively).Site mutation of nm23-H1 can affect the phosphorylation activity. H118 site was the key amino acid of kinase activity, P96 site maybe related to phosphotransferring, S120 was the site of histidine autophosphorylation and serine autophosphorylation, while the S44 site may be another amino acid which possessed NDPK activity.