Isolation and sequence determination of cDNA encoding T-protein of the glycine cleavage system.
T-protein is a component of the glycine cleavage system and catalyzes the tetrahydrofolate-dependent reaction. A partial cDNA for chicken T-protein was isolated by screening a chicken liver cDNA library with oligonucleotide probes based on amino acid sequences. The clone (CT5C) contains a 675-base pair insert encoding the C-terminal half of the T-protein. Screening of a bovine liver lambda gt10 cDNA library with the insert of CT5C as a probe detected a clone, BT5A, with an insert of about 2 kilobase pairs. The 1191-base pair coding region encodes a precursor protein of 397 amino acids (Mr 42,852) comprised of a 22-residue mitochondrial targeting sequence and a 375-residue mature protein (Mr 40,534). The 33 amino acids immediately following the targeting sequence correspond exactly to those determined by sequence analysis of the amino terminus of the purified bovine T-protein. The mature protein contains several hydrophilic segments with a cluster of arginine and lysine. The T-protein cDNA probe hybridized to an mRNA species of about 2 kilobases in bovine brain, lung, heart, and liver. A probe for H-protein hybridized with two species of mRNA in these tissues and weak signals were also found in spleen. Although the enzymatic activities of T-protein and H-protein were found in these tissues where transcripts were found, activity of P-protein was detected only in liver and brain. Southern blot analysis of genomic DNA suggested that T-protein is a single copy gene.
Okamura-Ikeda
K
K
Institute for Enzyme Research, University of Tokushima, Japan.
Fujiwara
K
K
Yamamoto
M
M
Hiraga
K
K
Motokawa
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002417:Cattle; D002645:Chickens; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D005998:Glycine / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014018:Tissue Distribution
D010750:Phosphoproteins; D010940:Plant Proteins; D012333:RNA, Messenger; C034921:T-substrate protein, Triticum aestivum; D004247:DNA; D005998:Glycine