A novel variant L263F in human inosine 5'-monophosphate dehydrogenase 2 is associated with diminished enzyme activity.
OBJECTIVE
Inosine 5'-monophosphate dehydrogenase 2 is required for purine synthesis in activated lymphocytes. Variants in the IMPDH2 gene may account for the large inter-individual variability in baseline enzyme activity, immunosuppressive efficacy and side effects in transplant recipients receiving mycophenolic acid. Therefore, the objective of this study was to identify and functionally characterize IMPDH2 variants.


METHODS
DNA samples from 152 solid organ transplant patients were screened at exons and exon/intron junctions of the IMPDH2 genes by PCR amplification followed by bidirectional direct DNA sequencing. Genetic variant was constructed by site-directed mutagenesis and transformed to an inosine 5'-monophosphate dehydrogenase-deficient strain of Escherichia coli h712. Proteins were purified to homogeneity and the enzymatic activity was measured by reduced nicotinamide adenine dinucleotide production.


RESULTS
Nine genetic variants were identified in the IMPDH2 gene, with frequencies of the rarer alleles ranging from 0.5 to 10.2%. A novel nonsynonymous variant L263F was identified, and the kinetic assay demonstrated that the inosine 5'-monophosphate dehydrogenase activity of L263F variant was decreased to 10% of the wild-type. The Ki for mycophenolic acid inhibition of the L263F variant was comparable with the wild-type, and the variant Km for inosine 5'-monophosphate and nicotinamide adenine dinucleotide did not change significantly.


CONCLUSIONS
IMPDH2 has low genetic diversity, but the nonsynonymous variant L263F has a significant impact on inosine 5'-monophosphate dehydrogenase activity. This novel functional variant may be one of the factors contributing to the inter-individual difference of baseline inosine 5'-monophosphate dehydrogenase activity as well as drug efficacy and adverse events in transplant patients.
Wang
Jian
J
Department of Pharmacy, University of Southern California, Los Angeles, California 90033, USA.
Zeevi
Adriana
A
Webber
Steve
S
Girnita
Diana M
DM
Addonizio
Linda
L
Selby
Rick
R
Hutchinson
Ian V
IV
Burckart
Gilbert J
GJ
D000483:Alleles; D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000235:genetics; D005091:Exons; D005787:Gene Frequency; D014644:Genetic Variation; D006801:Humans; D007168:IMP Dehydrogenase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism*; D007166:Immunosuppressive Agents / Q000494:pharmacology; D066298:In Vitro Techniques; D007438:Introns; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009173:Mycophenolic Acid / Q000494:pharmacology; D010597:Pharmacogenetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014181:Transplantation Immunology / Q000187:drug effects / Q000235:genetics
D007166:Immunosuppressive Agents; D011994:Recombinant Proteins; D004247:DNA; D007168:IMP Dehydrogenase; C501687:IMPDH2 protein, human; D009173:Mycophenolic Acid