Primary structure study of phosphofructokinase from Streptococcus lactis : a thesis presented [in] fulfilment of the requirement for the Master of Science in biochemistry at Massey University

Abstract

Phosphofructokinase is an important regulatory enzyme in the glycolytic pathway catalysing the phosphorylation of Fructose-6-phosphate to Fructose-1, 6-bisphosphate. Phosphofructokinase from Streptococcus lactis was isolated by chromatographic methods including DEAE Cellulose ion-exchange and Cibacron Blue Sepharose dye-ligand chromatographies. Phosphofructokinase from Streptococcus lactis was digested with trypsin, chymotrypsin, Staphylococcus aureus V8 protease and CNBr and the peptides obtained were sequenced and aligned against the sequences of Escherichia coli and Bacillus stearothermophilus phosphofructokinases. 326 out of 328 amino acid residues of Streptococcus lactis phosphofructokinase were obtained in this study. The comparison of Streptoccus lactis phosphofructokinase with Escherichia coli and Bacillus stearothermophilus phosphofructokinases showed that the sequence similarities among them are above 50%. Most of the secondary structures are conserved in Streptococcus lactis phosphofructokinase. The two 2-helices at the carboxyl terminal of bacterial phosphofructokinases are longer in Streptococcus lactis than in Escherichia coli and Bacillus stearothermophilus. The residues involved in binding of Fructose-6-phosphate/Fructose-1, 6-bisphosphate are the same in the bacterial phosphofructokinase and the residues involved in binding of ATP/ADP and binding of effectors have high degree of similarity.