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Functional expression of microbial phytase genes in industrial host
산업적 호스트를 이용한 미생물 유래의 파이테이즈 유전자의 기능적 발현에 관한 연구

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Authors
이재천
Advisor
최윤재
Issue Date
2003
Publisher
서울대학교 대학원
Keywords
파이테이즈Phytase대장균Pseudomonas효모E. coli단백질 발현PenicilliumPichia
Description
Thesis (master`s)--서울대학교 대학원 :농생명공학부,2003.
Abstract
Cereals, legumes, and oilseed crops are grown in over 90% of the world''s
harvested area. These crops serve as a major source of nutrients for humans and
animals. An important constituent in these crops is phytic acid(myo-inositol
hexaphosphate). the salt form, phytate, is the major storage form of phosphorus
and accounts for more than 80% of the total phosphorus in creals and legumes.
Phytase is an enzyme capable of hydrolyzing phytic acid to less-phosphorylated
myo-inositol derivates. Monogastric animals, such as pig, poultry and fish, are
not able to metabolized phytic acid, and therefore inorganic phosphate is added
to their diets to safety the phosphorus requirement. This consequently
contributes to phosphorus pollution problems in areas of intensive livestock
production.
Phytic acid also acts as an anti-nutritional agent in monogastric animals by
chelating various metal ions needed by the animal, such as calcium, copper, and
zinc. Therefore the enzymatic hydrolysis of phytic acid into less phosphorylated
myo-inositol derivatives in the intestine of monogastric animals is desirable.
Many attepts to enzymatically hydrolyze phytic acid have been made to improve
the nutitional value of feed and to decrease the amount of phosphorus excreted
by animals.
So, this study was conducted to construct the recombinant phytase gene from
Pseudomonas syringae MOK1 and Penicillium oxalicum PJ3 with commercial vectors
in industrial hosts and to characterize the recombinant proteins.
1. Psudomonas syringae MOK1 genes with pET vector were transfored into E. coli
BL21(DE3).
2. Transformed cells were induced by IPTG, after 6 h the cell extract were
assayed phytase activity. The result of phytase activity was about 0.4 U/ml
3. Penicillium oxalicum PJ3 phytase gene with pPICZ¥áA vector were transfored
into P. pastoris GS115 Mut+.
4. Recombinant phytase activity was increased steadily up to 120 h after the
methanol induction and cell growth pattern was also similar to increase ratio of
phytase activity.
5. The enzyme had a optimum pH of 4.5, and showed over 70% of the maximum
activity in the pH range of 3.7-5.0. The optimum temperature for the enzyme
activity was 55¡É, and showed over 70% of the maximum activity in the broad
temperature range of 38-61¡É.
6. The specific activity of the purified enzyme was 306.62U/mg of protein at
55¡É and pH 4.5.
7. The Km and Vmax values determined by the Line-Weaver Burk plot were 0.37 mM
and 526.3 U/mg respectively.
8. In SDS-PAGE analysis, the sample contains little minor contaminated proteins.
Upon deglycosylation in vitro by Endo H, the molecular size of the expressed
phytase by the control cells was reduced from 67kDa to 50kDa.
Language
English
URI
http://dcollection.snu.ac.kr:80/jsp/common/DcLoOrgPer.jsp?sItemId=000000057900

https://hdl.handle.net/10371/68517
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College of Agriculture and Life Sciences (농업생명과학대학)Dept. of Food and Animal Biotechnology (식품·동물생명공학부)Theses (Master's Degree_식품·동물생명공학부)
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