Protein Expression and Purification, Vol.83, No.2, 190-197, 2012
Molecular cloning, sequence analysis and expression in Escherichia coil of Camelus dromedarius glucose-6-phosphate dehydrogenase cDNA
This study determined the full length sequence of glucose-6-phosphate dehydrogenase cDNA (G6PD) from the Arabian camel Camelus dromedarius using reverse transcription polymerase chain reaction. The C. dromedarius G6PD has an open reading frame of 1545 bp, and the cDNA encodes a protein of 515 amino acid residues with a molecular weight of 59.0 KDa. The amino acid sequence showed the highest identity with Equus caballus (92%) and Homo sapiens (92%). The G6PD cDNA was cloned and expressed into Escherichia con as a fusion protein and was purified in a single chromatographic step using nickel affinity gel column. The purity and the molecular weight of the enzyme were checked on SDS-PAGE and the purified enzyme showed a single band on the gel with a molecular weight of 63.0 KDa. The specific activity of G6PD was determined to be 289.6 EU/mg protein with a fold purification of 95.45 and yield of 56.8%. (C) 2012 Elsevier Inc. All rights reserved.
Keywords:Arabian camel;Degenerate primers;Glucose-6-phosphate dehydrogenase;Reactive oxygen species (ROS)