Glutamine:fructose-6-phosphate amidotransferase(GFAT) is the rate-limiting
enzyme of the
hexosamine synthesis pathway. Products of this pathway have been implicated in
insulin resistance and
glucose toxicity. GFAT1 is ubiquitous, whereas GFAT2 is expressed mainly in the central nervous system. In the course of developing a competitive
reverse transcriptase-polymerase chain reaction assay, we noted that GFAT1
cDNA from muscle but not from other tissues migrated as a doublet. Subsequent cloning and sequencing revealed two GFAT1 mRNAs in both mouse and human skeletal muscles. The novel GFAT1
mRNA (GFAT1Alt [muscle selective variant of GFAT1]) is likely a splice variant. It is identical to GFAT1 except for a 48 or 54 bp insert in the mouse and human, respectively, at
nucleotide position 686 of the coding sequence, resulting in a 16 or 18
amino acid insert at position 229 of the
protein. GFAT1Alt is the predominant GFAT1
mRNA in mouse hindlimb muscle, is weakly expressed in the heart, and is undetectable in the brain, liver, kidney, lung, intestine, spleen, and 3T3-L1 adipocytes. In humans, it is strongly expressed in skeletal muscle but not in the brain. GFAT1 and GFAT1Alt expressed by recombinant
adenovirus infection in COS-7 cells displayed robust
enzyme activity and kinetic differences. The apparent K(m) of GFAT1Alt for
fructose-6-phosphate was approximately twofold higher than that of GFAT1, whereas K(i) for
UDP-
N-acetylglucosamine was approximately fivefold lower. Muscle
insulin resistance is a hallmark and predictor of
type 2 diabetes. Variations in the expression of GFAT
isoforms in muscle may contribute to predisposition to
insulin resistance.