Amino acids substitution to improve PhyFAUIA1 phytase activity for animal feed production

• Main Authors: Samsudin , Nurhusna, Mohd. Salleh, Hamzah, Noorbatcha, Ibrahim Ali, Amid, Azura
• Format: Article
• Language: English
• Published: Research Journal of Chemistry and Environment 2010
• Subjects: Q Science (General); TP248.13 Biotechnology
• Online Access: http://irep.iium.edu.my/2265/1/Pytase_Activity_328-333.pdf; http://irep.iium.edu.my/2265/; http://www.chemenviron.net

Phytate is largely unavailable to monogastric animal such as swine, poultry and fish, as they lack of sufficient endogenous enzymatic activity to hydrolyze phytate. The result is the elimination of precious nutrients that would be beneficial to their growth; furthermore they will excrete most of the indigestive phytate which can contribute to phosphorus being over applied to the land. Phosphorus has a beneficial impact on vegetative growth on land as well as marine vegetation, causing an increased growth of weeds. This enhanced vegetation consumes large amounts of oxygen, resulting in the loss of aquatic life and ultimately contributes to water pollution and eutrophication of ground water and aquatic environment. Phytase, a type of histidine acid phosphatase hydrolyzes phytin phosphorus and when present in an animal's digestive tract, benefits the animal while reducing total phosphorus levels in manure. Computer modelling is used to identify and examine the active site of phytase. The factors influencing the ligand binding strength in the active site is analyzed and computational site directed mutagenesis experiments were carried out to evaluate the effects of mutations on the binding strength before and after mutation. From the directive results of computational studies, point mutation was introduced by site directed mutagenesis using polymerase chain reaction (PCR). Mutagenesis is achieved by two step PCR procedure. Four primers were generated which two of them was design for carried the mutation at the point of interest which is complement to each other and the other two was design for unique restriction site. Several numbers of single, double as well as triple mutations has been introduced which will be further characterized to determine the activity of the enzyme as compared to the computational results.