| Marine yeast strains isolated from various marine habitats were studied from 2003 November to 2005 November. Isolation of yeasts strains were carried out under sterile conditions and were stored at -70°C at the laboratory of Molecular microbiology, UNESCO Chinese Center of Marine Biotechnology, Ocean University of China, Yushan Road, No. 5, China.During the experimental period, about 300 strains were isolated. Among them, a total of 177 isolates were classified to species level and 29 morphologically distinct yeast species were identified and genotyped. The highest species diversity was observed in sea water samples. Yarrowia lipolytica (39.8%) is the most common species identified yet host specific or habitat specific relationship was not clearly observed in this study. The complete YPD agar medium was used to store the yeast strains.Isolated strains were then screened for phytase activity using a modified minimal synthetic medium, which contained vitamins, minerals, salts, ammonium sulfate, agar and 0.5% sodium phytate as a sole source of carbon and phosphate. Strains with significant growth were selected and subjected for further enzyme assay. As a result ten strains exhibited comparatively high enzyme activity and Yarrowia lipolytica (W2B) and Kodamea ohmeri (BG3) had the highest extra-cellular phytase activity. Strain YF 12C indicated the highest cell bound enzyme activity and this strain was screened from the gut of Sea cucumber (Holothuria scabra). Interestingly, the optimum pH and temperature for crude enzyme of all strains were between 5 - 9 and 50°C– 65°C respectively.Among these ten isolates, WZ1 was from the gut of fish (Scomberomorus niphonius), W2B and YF12C from sea cucumber (Holothuria scabra), whereas BG3, YF04C and YF08 were from the gut of fish species (Hexagrammes otakii and Synecogobius hasts). The sea water isolates were N12C and MA6 including NY4E and MB2 from Salterns and Southern sea of China respectively. The results of this study revealed that the predominant phytase secreting strains were the ones isolated from the gut of marine animals. In addition, this study indicated the diverse habitats of these strains. To date, few terrestrial yeasts species have been reported as phytase producers but marine yeasts were in scarce as we used in this research.Ten strains with relatively high phytase activity were then characterized by genetic, physiological and biochemical analyses to clarify their taxonomic position. The 18S rDNA partial genomic sequences and ITS sequences of the yeast strains were examined and analyzed for similarity comparison using NCBI BLAST program and deposited at NCBI data bank. Phylogenetic tree was constructed by using PHILIP program. Taxonomic studies confirmed that strains WZ1, W2B, BG3, YF04C, and YF08 were similar to Hanseniaspora uvarum, Yarrowia lipolytica, Kodamaea ohmeri, Issatchenkia orientalis and Yarrowia lipolytica, respectively, while both N12C and NY4E to Candida carpophila, and MA6, YF12C and MB2 were similar to Candida tropicalis, respectively.In the current study, yeast strain Hanseniaspora uvarum - WZ1 was selected for purification and was grown for four days at 28°C in optimized medium. Extra-cellular phytase from was purified using gel filtration chromatography column of Sephadex TM G-100. The optimum pH and temperature of the purified enzyme was pH 5 and 60°C, respectively. These values were different from that of the crude enzyme. The molecular weight was about 46.3 kDa. Purified phytase was not inhibited by CaCl2, MgCl2, NH4Cl and ZnCl2 suggesting that Ca+2, Mg+2 NH4+ and Zn+2 ions are not essential for activity. The enzyme was strongly inhibited by Cu +2 however, Co +, Li + and Mn +2 were less effective. The Km and V max values were 0.392 mmol and 3.889 mmol/min.
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