ABSTRACT: Tenofovir (TFV) is a nucleotide reverse transcriptase inhibitor prescribed for the treatment and prevention of human immunodeficiency virus infection, and the treatment of chronic hepatitis B virus infection. Here, we demonstrate that creatine kinase brain-type (CKB) can form tenofovir-diphosphate (TFV-DP), the pharmacologically active metabolite, in vitro, and identify nine missense mutations (C74S, R96P, S128R, R132H, R172P, R236Q, C283S, R292Q, and H296R) that diminish this activity. Additional characterization of these mutations reveal that five (R96P, R132H, R236Q, C283S, and R292Q) have ATP dephosphorylation catalytic efficiencies less than 20% of wild-type (WT), and seven (C74S, R96P, R132H, R172P, R236Q, C283S, and H296P) induce thermal instabilities. To determine the extent CKB co
