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Ghatge MS, Karve SS, David TM, et al. Inactive mutants of human pyridoxine 5'-phosphate oxidase: a possible role for a noncatalytic pyridoxal 5'-phosphate tight binding site. FEBS Open Bio. 2016;6(5):398-408doi: 10.1002/2211-5463.12042.
Ghatge, M. S., Karve, S. S., David, T. M., Ahmed, M. H., Musayev, F. N., Cunningham, K., Schirch, V., & Safo, M. K. (2016). Inactive mutants of human pyridoxine 5'-phosphate oxidase: a possible role for a noncatalytic pyridoxal 5'-phosphate tight binding site. FEBS open bio, 6(5), 398-408. https://doi.org/10.1002/2211-5463.12042
Ghatge, Mohini S, et al. "Inactive mutants of human pyridoxine 5'-phosphate oxidase: a possible role for a noncatalytic pyridoxal 5'-phosphate tight binding site." FEBS open bio vol. 6,5 (2016): 398-408. doi: https://doi.org/10.1002/2211-5463.12042
Ghatge MS, Karve SS, David TM, Ahmed MH, Musayev FN, Cunningham K, Schirch V, Safo MK. Inactive mutants of human pyridoxine 5'-phosphate oxidase: a possible role for a noncatalytic pyridoxal 5'-phosphate tight binding site. FEBS Open Bio. 2016 Mar 22;6(5):398-408. doi: 10.1002/2211-5463.12042. eCollection 2016 May. PMID: 27419045; PMCID: PMC4856418.
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FEBS Open Bio. 2016 Mar 22;6(5):398-408. doi: 10.1002/2211-5463.12042. eCollection 2016 May.

Inactive mutants of human pyridoxine 5'-phosphate oxidase: a possible role for a noncatalytic pyridoxal 5'-phosphate tight binding site.

FEBS open bio

Mohini S Ghatge, Sayali S Karve, Tanya M S David, Mostafa H Ahmed, Faik N Musayev, Kendra Cunningham, Verne Schirch, Martin K Safo

Affiliations

  1. Department of Medicinal Chemistry School of Pharmacy and Institute for Structural Biology, Drug Discovery and Development Virginia Commonwealth University Richmond VA USA.

PMID: 27419045 PMCID: PMC4856418 DOI: 10.1002/2211-5463.12042

Abstract

Pyridoxal 5'-phosphate (PLP) is a cofactor for many vitamin B6-requiring enzymes that are important for the synthesis of neurotransmitters. Pyridoxine 5'-phosphate oxidase (PNPO) is one of two enzymes that produce PLP. Some 16 known mutations in human PNPO (hPNPO), including R95C and R229W, lead to deficiency of PLP in the cell and have been shown to cause neonatal epileptic encephalopathy (NEE). This disorder has no effective treatment, and is often fatal unless treated with PLP. In this study, we show that R95C hPNPO exhibits a 15-fold reduction in affinity for the FMN cofactor, a 71-fold decrease in affinity for the substrate PNP, a 4.9-fold decrease in specific activity, and a 343-fold reduction in catalytic activity, compared to the wild-type enzyme. We have reported similar findings for R229W hPNPO. This report also shows that wild-type, R95C and R229W hPNPO bind PLP tightly at a noncatalytic site and transfer it to activate an apo-B6 enzyme into the catalytically active holo-form. We also show for the first time that hPNPO forms specific interactions with several B6 enzymes with dissociation constants ranging from 0.3 to 12.3 μm. Our results suggest a possible in vivo role for the tight binding of PLP in hPNPO, whether wild-type or variant, by protecting the very reactive PLP, and transferring this PLP directly to activate apo-B6 enzymes.

Keywords: enzyme mutation; neonatal epileptic encephalopathy; neurotransmitters; pyridoxal 5′‐phosphate; pyridoxine 5′‐phosphate oxidase; vitamin B6

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