Cloning and characterization of an alternative transcript of ovine glucose 6-phosphate dehydrogenase gene: comparative approach between ruminant and non-ruminant species.
Glucose 6-phosphate dehydrogenase (G6PD) plays an important role in ruminant's lipogenesis, as it provides necessary compounds of NADPH for the synthesis of fatty acids catalyzing the first committed reaction in the pentose phosphate pathway. In this work the full length ovine glucose 6-phosphate dehydrogenase cDNA was isolated using a polymerase chain reaction based strategy. Two isoforms (OG6PDA and OG6PDB) were detected encoding a protein of 515 and 524 amino acids, respectively. Both deduced amino acid sequences reveal a well conserved protein containing all the important residues for its catalytic role. The extra nine amino acids encoded by OG6PDB cause a frameshift in the polypeptide chain resulting in changes around the area of the potential substrate binding site. A three-dimensional model of ovine G6PD protein shows that this frameshift cause structural changes in the catalytic binding "pocket" of the molecule. Southern blot and RT analysis revealed that ovine G6PD appears as a single copy gene while it is expressed, with slight variability, in all tissues analyzed. Moreover, expression analysis of the ovine G6PD isoforms showed that OG6PDB is expressed only in tissues where lipogenesis is high in ruminants. Thus, we hypothesize that in ruminants G6PD may be regulated by the ratio of the two transcripts, according to the existence stimulus.
Laliotis
George P
GP
Department of Animal Science, Laboratory of Animal Breeding and Husbandry, Agricultural University of Athens, Iera Odos 75, 118 55 Athens, Greece.
Argyrokastritis
Alexandros
A
Bizelis
Iosif
I
Rogdakis
Emmanuel
E
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D020134:Catalytic Domain / Q000235:genetics; D003001:Cloning, Molecular; D004247:DNA / Q000737:chemistry / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling; D005954:Glucosephosphate Dehydrogenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008297:Male; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012418:Ruminants / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012756:Sheep / Q000235:genetics*; D013045:Species Specificity
D018076:DNA, Complementary; D007527:Isoenzymes; D004247:DNA; D005954:Glucosephosphate Dehydrogenase