Exploiting AGPase genes and encoded proteins to prioritize development of optimum engineered strains in microalgae towards sustainable biofuel production.

Although ADP glucose pyrophosphorylase (AGPase), with two large subunits (ls) and two small subunits (ss), is a promising knockout target for increasing the neutral lipid content, the details regarding the sequence-structure features and their distribution within metabolic system in microalgae is rather limited. Against this backdrop, a comprehensive genome-wide comparative analysis on 14 sequenced microalgal genomes was performed. For the first time the heterotetrameric structure of the enzyme and the interaction of the catalytic unit with the substrate was also studied. Novel findings of the present study includes: (i) at the DNA level, the genes controlling the ss are more conserved than those controlling the ls; the variation in both the gene groups is mainly due to exon number, exon length and exon phase distribution; (ii) at protein level, the ss genes are more conserved relative to those for ls; (III) three putative key consensus sequences 'LGGGAGTRLYPLTKNRAKPAV', 'WFQGTADAV' and 'ASMGIYVFRKD' were ubiquitously conserved in all the AGPases; (iv) molecular dynamics investigations revealed that the modeled AGPase heterotetrameric structure, from oleaginous algae Chlamydomonas reinharditii, was completely stable in real time environment; (v) The binding interfaces of catalytic unit, ssAGPase, from C. reinharditii with α-D-glucose 1-phosphate (αGP) was also analyzed. The results of the present study have provided system-based insights into the structure-function of the genes and encoded proteins, which provided clues for exploitation of variability in these genes that, could be further utilized to design site-specific mutagenic experiments for engineering of microalgal strains towards sustainable development of biofuel.