| Summary: | 碩士 === 國立屏東科技大學 === 獸醫學系 === 93 === Yeast cell containing a great deal of high cost nontoxic enzymes, proteins etc. provides a good source for further economic utilization. In this study, The Saccharomyces cerevisia ( SC ) of the Angel baking powder seed yeast ( SC04 ) gave the best growth in variant commercial and self-prepared medium for cultivating SC was chosed from 11 commercial baking powder seeds for further mass production. Futhermore, Alcohol dehydrogenase ( ADH ) were extracted from a complete mechanically breaking up SC cells with distilled water ( DW ), then purified with cold acetone and saturated ammonium sulfate, and aldehyde dehydrogenase ( ALDH ) was then purified from the above DW extract, then further purified with pooled acid and protoamine solution. A 33210 units of ADH ( 47 kDa ) and 97 units of ALDH ( 52 kDa ) were harvested from the powder per gram as measuring by the spectrophotometer detecting the nicotinamide adenine dinucleotide ( NADH ) formation under 340 nm ultraviolet light. The whole length of ADH ( 1062 bp ) and ALDH ( 1536 bp ) gene were polymerase chain reaction ( PCR ) amplified and inserted in the pET32a vector individually. The recombinant plasmid was transformated into E. coli BL21 expression system. A recombinant ADH ( 59 kDa ) or ALDH ( 76 kDa ) was expressed from each system with Western blotting identified, but without providing any dehydrogenation activity to ethyl alcohol or acetoaldehyde. ADH and ALDH expressed by the E. coli expression system as done in this experiment is not able to produce the product with good enzymatic activity as the traditional extraction from the baking yeast does.
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