TY - JOUR
T1 - Naturally occurring mutations to muscle-type creatine kinase impact its canonical and pharmacological activities in a substrate-dependent manner in vitro
AU - Mosher, Eric P.
AU - Eberhard, Colten D.
AU - Bumpus, Namandje N.
N1 - Funding Information:
This work was supported by National Institutes of Health (NIH) National Institute of Allergy and Infectious Diseases [Grant R01AI128781] and National Institute on Aging [Grant R01AG064908] (to N.N.B.) and a 2020 PhRMA Foundation Predoctoral Fellowship in Pharmacology and Toxicology awarded to E.P.M. E.P.M. was also supported by NIH National Institute of General Medical Sciences [Grant T32-GM008763]. The authors report no conflict of interest. dx.doi.org/10.1124/molpharm.121.000348.
Publisher Copyright:
© 2021 by The Author(s)
PY - 2021/12/1
Y1 - 2021/12/1
N2 - Tenofovir (TFV) is a key component of human immunodeficiency virus (HIV) pre-exposure prophylaxis (PrEP). TFV is a nucleotide analog reverse-transcriptase inhibitor prodrug that requires two separate phosphorylation reactions by intracellular kinases to form the active metabolite tenofovir-diphosphate (TFV-DP). Muscle-type creatine kinase (CKM) has previously been demonstrated to be the kinase most responsible for the phosphorylation of tenofovir-mono-phosphate (TFV-MP) to the active metabolite in colon tissue. Because of the importance of CKM in TFV activation, genetic variation in CKM may contribute to interindividual variability in TFV-DP levels. In the present study, we report 10 naturally occurring CKM mutations that reduced TFV-MP phosphorylation in vitro: T35I, R43Q, I92M, H97Y, R130H, R132C, F169L, Y173C, W211R, V280L, and N286I. Interestingly, of these 10, only 4—R130H, R132C, W211R, and N286I—reduced both canonical CKM activities: ADP phosphorylation and ATP dephosphorylation. Although positions 130, 132, and 286 are located in the active site, the other mutations that resulted in decreased TFV-MP phosphorylation occur elsewhere in the protein structure. Four of these eight mutations—T35I, R43Q, I92M, and W211R—were found to decrease the thermal stability of the protein. Additionally, the W211R mutation was found to impact protein structure both locally and at a distance. These data suggest a substrate-specific effect such that certain mutations are tolerated for canonical activities while being deleterious toward the pharmacological activity of TFV activation, which could influence PrEP outcomes.
AB - Tenofovir (TFV) is a key component of human immunodeficiency virus (HIV) pre-exposure prophylaxis (PrEP). TFV is a nucleotide analog reverse-transcriptase inhibitor prodrug that requires two separate phosphorylation reactions by intracellular kinases to form the active metabolite tenofovir-diphosphate (TFV-DP). Muscle-type creatine kinase (CKM) has previously been demonstrated to be the kinase most responsible for the phosphorylation of tenofovir-mono-phosphate (TFV-MP) to the active metabolite in colon tissue. Because of the importance of CKM in TFV activation, genetic variation in CKM may contribute to interindividual variability in TFV-DP levels. In the present study, we report 10 naturally occurring CKM mutations that reduced TFV-MP phosphorylation in vitro: T35I, R43Q, I92M, H97Y, R130H, R132C, F169L, Y173C, W211R, V280L, and N286I. Interestingly, of these 10, only 4—R130H, R132C, W211R, and N286I—reduced both canonical CKM activities: ADP phosphorylation and ATP dephosphorylation. Although positions 130, 132, and 286 are located in the active site, the other mutations that resulted in decreased TFV-MP phosphorylation occur elsewhere in the protein structure. Four of these eight mutations—T35I, R43Q, I92M, and W211R—were found to decrease the thermal stability of the protein. Additionally, the W211R mutation was found to impact protein structure both locally and at a distance. These data suggest a substrate-specific effect such that certain mutations are tolerated for canonical activities while being deleterious toward the pharmacological activity of TFV activation, which could influence PrEP outcomes.
UR - http://www.scopus.com/inward/record.url?scp=85120703898&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85120703898&partnerID=8YFLogxK
U2 - 10.1124/molpharm.121.000348
DO - 10.1124/molpharm.121.000348
M3 - Article
C2 - 34561299
AN - SCOPUS:85120703898
SN - 0026-895X
VL - 100
SP - 588
EP - 596
JO - Molecular Pharmacology
JF - Molecular Pharmacology
IS - 6
ER -