Elevated homocysteine (Hcy) in the blood is an established risk factor for cardiovascular disease. Increases in dietary folate and B6 have been shown to lower Hcy levels. There are however, certain groups with other health problems where Hcy remains elevated, these include heart transplant recipients, diabetics, women with preclampsia or retarded fetal growth, end stage renal disease and Parkinson's disease.
The aim of this proposal is to elucidate how nutritional insufficiency of folate and B6 affect the pathways of Hcy metabolism in mammalian cells. There are four specific aims: (1) the development of rapid enzyme-based assays for 5,10-methyleneTHF, B6 vitamers and homocysteine; (2) to determine the direction of flux of 1-carbon (1-C) groups in the cytosol and mitochondria of cells in culture, with special emphasis on serine hydroxmethyltransferase (SHMT); (3) to determine the role of mitochondria in the supply of 1-C groups to the cytosol; and (4) to determine the relationship of folate pools and metabolic levels of homocysteine with several different cell lines when either folate or B6 are limiting growth factors. Three hypotheses will be tested, which are: (1) that the role of cytosolic SHMT is not to generate 1-C units but to regulate the levels of glycine and 5,10-methyleneTHF in the cytosol; (2) that 1-C groups used by the cytosol are generated by the mitochondria as formate; and (3) Hcy levels are related to the level of 5,10-methyleneTHF in the cytosol.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
1R01DK056648-01
Application #
6027216
Study Section
Nutrition Study Section (NTN)
Program Officer
May, Michael K
Project Start
2000-05-01
Project End
2005-01-31
Budget Start
2000-05-01
Budget End
2001-01-31
Support Year
1
Fiscal Year
2000
Total Cost
$171,249
Indirect Cost
Name
Virginia Commonwealth University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
City
Richmond
State
VA
Country
United States
Zip Code
23298
Safo, Martin K; Musayev, Faik N; di Salvo, Martino L et al. (2006) Crystal structure of pyridoxal kinase from the Escherichia coli pdxK gene: implications for the classification of pyridoxal kinases. J Bacteriol 188:4542-52
Schirch, Verne; Szebenyi, Doletha Me (2005) Serine hydroxymethyltransferase revisited. Curr Opin Chem Biol 9:482-7
Fu, Tzu-Fun; Hunt, Sharyn; Schirch, Verne et al. (2005) Properties of human and rabbit cytosolic serine hydroxymethyltransferase are changed by single nucleotide polymorphic mutations. Arch Biochem Biophys 442:92-101
Safo, Martin K; Musayev, Faik N; Hunt, Sharyn et al. (2004) Crystal structure of the PdxY Protein from Escherichia coli. J Bacteriol 186:8074-82
Szebenyi, Doletha M E; Musayev, Faik N; di Salvo, Martino L et al. (2004) Serine hydroxymethyltransferase: role of glu75 and evidence that serine is cleaved by a retroaldol mechanism. Biochemistry 43:6865-76
di Salvo, Martino L; Safo, Martin K; Musayev, Faik N et al. (2003) Structure and mechanism of Escherichia coli pyridoxine 5'-phosphate oxidase. Biochim Biophys Acta 1647:76-82
Fu, T F; Rife, J P; Schirch, V (2001) The role of serine hydroxymethyltransferase isozymes in one-carbon metabolism in MCF-7 cells as determined by (13)C NMR. Arch Biochem Biophys 393:42-50