Isolation, characterization, and expression of a cDNA encoding N-acetylglucosaminyltransferase V.
A cDNA clone for the complete coding sequence for alpha-1,3(6)-mannosylglycoprotein beta-1,6-N-acetylglucosaminyltransferase V (GlcNAc-T V, EC 2.4.1.155) was isolated and expressed in COS-7 cells. Degenerate oligonucleotide primers for polymerase chain reaction were synthesized based on the amino acid sequence of three tryptic peptides isolated from affinity-purified GlcNAc-T V. Polymerase chain reaction amplimers were isolated from rat and mouse mRNA. A cDNA-encoding full-length enzyme was isolated from a rat 1 cell (EJ-ras-transformed) library and sequenced. Transient expression of this clone in COS-7 cells, followed by enzymatic activity assays, demonstrated that this cDNA sequence encodes GlcNAc-T V. Northern analysis of rat kidney mRNA revealed a single band corresponding to a length of about 7 kilobases. Sequence analysis of the cDNA clone demonstrated an open reading frame that encoded a type II membrane protein of 740 amino acids.
Shoreibah
M
M
Department of Biochemistry, University of Georgia, Athens 30602.
Perng
G S
GS
Adler
B
B
Weinstein
J
J
Basu
R
R
Cupples
R
R
Wen
D
D
Browne
J K
JK
Buckhaults
P
P
Fregien
N
N
Pierce
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000302:isolation & purification; D007668:Kidney / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats
D012333:RNA, Messenger; D004247:DNA; D017351:N-Acetylglucosaminyltransferases; C052254:alpha-1,6-mannosylglycoprotein beta 1,6-N-acetylglucosaminyltransferase