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Human mitochondrial C1-tetrahydrofolate synthase: gene structure, tissue distribution of the mRNA, and immunolocalization in Chinese hamster ovary calls.
J Biol Chem. 2003 Oct 31; 278(44):43178-43187.JB

Abstract

C1-tetrahydrofolate (THF) synthase is a trifunctional enzyme found in eukaryotes that contains the activities 10-formyl-THF synthetase, 5,10-methenyl-THF cyclohydrolase, and 5,10-methylene-THF dehydrogenase. The cytoplasmic isozyme of C1-THF synthase is well characterized in a number of mammals, including humans; but a mitochondrial isozyme has been previously identified only in the yeast Saccharomyces. Here, we report the identification and characterization of the human gene encoding a functional mitochondrial C1-THF synthase. The gene spans 236 kilobase pairs on chromosome 6 and consists of 28 exons plus one alternative exon. The gene encodes a protein of 978 amino acids, including an N-terminal mitochondrial targeting sequence. The mitochondrial isozyme is 61% identical to the human cytoplasmic isozyme. Expression of the gene was detected in most human tissues, but transcripts were highest in placenta, thymus, and brain. Two mRNAs were detected, a 3.6-kb transcript and a 1.1-kb transcript, and both transcripts were observed in varying ratios in each tissue. The shorter transcript results from an alternative splicing event, where exon 7 is spliced to exon 8a instead of exon 8. Exon 8a is derived from an exonized Alu sequence, sharing no homology with exon 8 of the long transcript, and encodes just 15 amino acids followed by a stop codon and a polyadenylation signal. This short transcript potentially encodes a bifunctional enzyme lacking 10-formyl-THF synthetase activity. Both transcripts initiate at the same 5'-site, 107 nucleotides up-stream of the ATG start codon. The full-length (2934 bp) cDNA fused to a C-terminal V5 epitope tag was expressed in Chinese hamster ovary cells. Immunoblots of subfractionated cells revealed a 107-kDa protein only in the mitochondrial fractions of these cells, confirming the mitochondrial localization of the protein. Yeast cells expressing the full-length human cDNA exhibited elevated 10-formyl-THF synthetase activity, confirming its identification as the human mitochondrial C1-THF synthase.

Authors+Show Affiliations

Department of Chemistry & Biochemistry, Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin TX 78712.Department of Chemistry & Biochemistry, Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin TX 78712.Department of Nutritional Sciences, University of California, Berkeley CA 94720.Department of Nutritional Sciences, University of California, Berkeley CA 94720.Department of Chemistry & Biochemistry, Institute for Cellular and Molecular Biology, The University of Texas at Austin, Austin TX 78712.

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

12937168

Citation

Prasannan, Priya, et al. "Human Mitochondrial C1-tetrahydrofolate Synthase: Gene Structure, Tissue Distribution of the mRNA, and Immunolocalization in Chinese Hamster Ovary Calls." The Journal of Biological Chemistry, vol. 278, no. 44, 2003, pp. 43178-43187.
Prasannan P, Pike S, Peng K, et al. Human mitochondrial C1-tetrahydrofolate synthase: gene structure, tissue distribution of the mRNA, and immunolocalization in Chinese hamster ovary calls. J Biol Chem. 2003;278(44):43178-43187.
Prasannan, P., Pike, S., Peng, K., Shane, B., & Appling, D. R. (2003). Human mitochondrial C1-tetrahydrofolate synthase: gene structure, tissue distribution of the mRNA, and immunolocalization in Chinese hamster ovary calls. The Journal of Biological Chemistry, 278(44), 43178-43187. https://doi.org/10.1074/jbc.M304319200
Prasannan P, et al. Human Mitochondrial C1-tetrahydrofolate Synthase: Gene Structure, Tissue Distribution of the mRNA, and Immunolocalization in Chinese Hamster Ovary Calls. J Biol Chem. 2003 Oct 31;278(44):43178-43187. PubMed PMID: 12937168.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Human mitochondrial C1-tetrahydrofolate synthase: gene structure, tissue distribution of the mRNA, and immunolocalization in Chinese hamster ovary calls. AU - Prasannan,Priya, AU - Pike,Schuyler, AU - Peng,Kun, AU - Shane,Barry, AU - Appling,Dean R, Y1 - 2003/08/22/ PY - 2003/8/26/pubmed PY - 2003/12/25/medline PY - 2003/8/26/entrez SP - 43178 EP - 43187 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 278 IS - 44 N2 - C1-tetrahydrofolate (THF) synthase is a trifunctional enzyme found in eukaryotes that contains the activities 10-formyl-THF synthetase, 5,10-methenyl-THF cyclohydrolase, and 5,10-methylene-THF dehydrogenase. The cytoplasmic isozyme of C1-THF synthase is well characterized in a number of mammals, including humans; but a mitochondrial isozyme has been previously identified only in the yeast Saccharomyces. Here, we report the identification and characterization of the human gene encoding a functional mitochondrial C1-THF synthase. The gene spans 236 kilobase pairs on chromosome 6 and consists of 28 exons plus one alternative exon. The gene encodes a protein of 978 amino acids, including an N-terminal mitochondrial targeting sequence. The mitochondrial isozyme is 61% identical to the human cytoplasmic isozyme. Expression of the gene was detected in most human tissues, but transcripts were highest in placenta, thymus, and brain. Two mRNAs were detected, a 3.6-kb transcript and a 1.1-kb transcript, and both transcripts were observed in varying ratios in each tissue. The shorter transcript results from an alternative splicing event, where exon 7 is spliced to exon 8a instead of exon 8. Exon 8a is derived from an exonized Alu sequence, sharing no homology with exon 8 of the long transcript, and encodes just 15 amino acids followed by a stop codon and a polyadenylation signal. This short transcript potentially encodes a bifunctional enzyme lacking 10-formyl-THF synthetase activity. Both transcripts initiate at the same 5'-site, 107 nucleotides up-stream of the ATG start codon. The full-length (2934 bp) cDNA fused to a C-terminal V5 epitope tag was expressed in Chinese hamster ovary cells. Immunoblots of subfractionated cells revealed a 107-kDa protein only in the mitochondrial fractions of these cells, confirming the mitochondrial localization of the protein. Yeast cells expressing the full-length human cDNA exhibited elevated 10-formyl-THF synthetase activity, confirming its identification as the human mitochondrial C1-THF synthase. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/12937168/Human_mitochondrial_C1_tetrahydrofolate_synthase:_gene_structure_tissue_distribution_of_the_mRNA_and_immunolocalization_in_Chinese_hamster_ovary_calls_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=12937168 DB - PRIME DP - Unbound Medicine ER -