3l3r
From Proteopedia
Structural, Computational and Kinetic Data for Antifolate Interactions Against Pneumocystis jirovecii, Pneumocystis carinii and Human Dihydrofolate Reductase and Their Active Site Mutants
Structural highlights
Disease[DYR_HUMAN] Defects in DHFR are the cause of megaloblastic anemia due to dihydrofolate reductase deficiency (DHFRD) [MIM:613839]. DHFRD is an inborn error of metabolism, characterized by megaloblastic anemia and/or pancytopenia, severe cerebral folate deficiency, and cerebral tetrahydrobiopterin deficiency. Clinical features include variable neurologic symptoms, ranging from severe developmental delay and generalized seizures in infancy, to childhood absence epilepsy with learning difficulties, to lack of symptoms.[1] [2] Function[DYR_HUMAN] Key enzyme in folate metabolism. Contributes to the de novo mitochondrial thymidylate biosynthesis pathway. Catalyzes an essential reaction for de novo glycine and purine synthesis, and for DNA precursor synthesis. Binds its own mRNA and that of DHFRL1.[3] [4] Publication Abstract from PubMedA major concern of immunocompromised patients, in particular those with AIDS, is susceptibility to infection caused by opportunistic pathogens such as Pneumocystis jirovecii, which is a leading cause of pneumonia in immunocompromised patients. We report the first kinetic and structural data for 2,4-diamino-6-[(2',5'-dichloro anilino)methyl]pyrido[2,3-d]pyrimidine (OAAG324), a potent inhibitor of dihydrofolate reductase (DHFR) from P. jirovecii (pjDHFR), and also for trimethoprim (TMP) and methotrexate (MTX) with pjDHFR, Pneumocystis carinii DHFR (pcDHFR), and human DHFR (hDHFR). OAAG324 shows a 9.0-fold selectivity for pjDHFR (Ki, 2.7 nM) compared to its selectivity for hDHFR (Ki, 24.4 nM), whereas there is only a 2.3-fold selectivity for pcDHFR (Ki, 6.3 nM). In order to understand the determinants of inhibitory potency, active-site mutations of pj-, pc-, and hDHFR were explored to make these enzymes more like each other. The most unexpected observations were that the variant pcDHFR forms with K37Q and K37Q/F69N mutations, which made the enzyme more like the human form, also made these enzymes more sensitive to the inhibitory activity of OAAG324, with Ki values of 0.26 and 0.71 nM, respectively. A similar gain in sensitivity was also observed for the hDHFR N64F variant, which showed a lower Ki value (0.58 nM) than native hDHFR, pcDHFR, or pjDHFR. Structural data are reported for complexes of OAAG324 with hDHFR and its Q35K and Q35S/N64F variants and for the complex of the K37S/F69N variant of pcDHFR with TMP. These results provide useful insight into the role of these residues in the optimization of highly selective inhibitors of DHFR against the opportunistic pathogen P. jirovecii. Kinetic and Structural Analysis for Potent Antifolate Inhibition of Pneumocystis jirovecii, Pneumocystis carinii, and Human Dihydrofolate Reductases and Their Active-Site Variants.,Cody V, Pace J, Queener SF, Adair OO, Gangjee A Antimicrob Agents Chemother. 2013 Jun;57(6):2669-77. doi: 10.1128/AAC.00172-13., Epub 2013 Apr 1. PMID:23545530[5] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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