Expression and characterization of Guamerin, an elastase inhibitor in Escherichia coli = 대장균에서 일라스테즈 억제제인 거머린의 발현과, 재조합 거머린의 특성에 관한 연구

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Guamerin, a cysteine-rich protein from the Korean leech Hirudo nipponia, is a potent inhibitor of elastase. The guamerin encoding gene was obtained by polymerase chain reaction and expressed in Escherichia coli under the control of T7lac promoter by using a pET22b vector, which has a pelB leader sequence and a 6 X Histidine tag at the N-terminal and C-terminal regions, respectively. Recombinant guamerin was purified using His$\cdot$Bind affinity column and reverse-phase high pressure liquid chromatography. N-terminal amino acid sequence analysis, compostion analysis and electrospray ionization mass spectrometry confirmed that recombinant guamerin had correctly processed N-terminus, expected amino acid sequence and molecular mass. The activity of recombinat guamerin was very similar to that of native protein. Double reciprocal plotting showed that the recombinant guamerin had inhibition constant of $8.8 \times 10^{-9}$ M against elastase, which is comparable to native guamerin. Recombinant guamerin formed a protease-inhibitor complex which was demonstrated by gel permeation chromatography. The recombinant guamerin may provide a useful tool for the evaluation of the elastase inhibitory mechanism and for treating various inflammatory human diseases .
Advisors
Kang, Ke-Won강계원
Description
한국과학기술원 : 생물과학과,
Publisher
한국과학기술원
Issue Date
1999
Identifier
156319/325007 / 000973015
Language
eng
Description

학위논문(석사) - 한국과학기술원 : 생물과학과, 1999.2, [ v, 37 p. ]

Keywords

Expression; Inhibitor; Elastase; Leech; Antistasin; 안티스타신; 발현; 억제제; 일라스테즈; 거머리

URI
http://hdl.handle.net/10203/27889
Link
http://library.kaist.ac.kr/search/detail/view.do?bibCtrlNo=156319&flag=dissertation
Appears in Collection
BS-Theses_Master(석사논문)
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