Screening, extraction and purification for tannase produced from Iraqi Klebsiella pneumonia isolates and molecular detection of tanA gene

In this study, tannase enzyme was produced from an Iraq Klebsiella pneumoniae isolates. K. pneumoniae were identified by Vitek system and confirmed by housekeeping 16s rRNA gene (amplified size 155 bp). Tannase was genotpically detected by amplification tanA gene (amplified size 210 bp) followed by sequencing. The tannase activity reached its maximum level when this isolate was cultivated under the optimal conditions, which is consisted of using 2.8 g of nutrient agar containing 2% (w/v) tannic acid as a carbon source at pH 5.5 and temperature of 37°C for 24 h. The Tannase had been purified by using three methods: ammonium sulphate, ion exchange and gel filtration. The first method leads to gain a tannase precipitation at 70% ammonium sulphate which is considered as a partial purification where tannase activity was 80U/ml. In comparison, 300 U/mg tannase activity had been gained by using ion exchange with 4.31 fold of purification and a yield of 21.4%. Finally, a tannase activity of 500 U/mg is gained by using gel filtration with 5.75 fold of purification and a yield of 21.4%. The purified tannase is a single peptide with approximate molecular mass of 46.5 kDa as assessed by SDS-PAGE.