Human mitochondrial NAD-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase was expressed in a soluble and active form in Escherichia coli, purified to homogeneity for the first time and characterized. Phosphate is absolutely required for the dehydrogenase activity. NADP can be utilized by the dehydrogenase with a low turnover rate. The properties of the enzyme suggest that it has evolved from NADP to NAD specificity by the unique mechanism of substituting inorganic phosphate for the 2$ sp prime$-phosphate of NADP. The role of Mg$ sp{++}$ in this process is not understood, but it is required for activity with both dinucleotides. This enzyme is proposed to be a mammalian homolog of the product of the yeast MIS1 gene, functioning in providing formyltetrahydrofolate for the synthesis of formylmethionyl tRNA required for the initiation of protein synthesis in mitochondria. The two activities of this bifunctional enzyme are resolved significantly by diethylpyrocarbonate modification of at least two histidines. Most striking is the ability to activate the cyclohydrolase up to 50% independently of the dehydrogenase by modification with diethylpyrocarbonate in the presence of ligands: Mg$ sp{++}/$phosphate/NAD/folate.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.41792 |
| Date | January 1993 |
| Creators | Yang, Xiao-Ming |
| Contributors | MacKenzie, Robert E. (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Biochemistry.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001397021, proquestno: NN94725, Theses scanned by UMI/ProQuest. |
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