| Summary: | 碩士 === 國立臺灣大學 === 生化科學研究所 === 86 === Nephrosin is a newly discovered member of the astacin family, a group
of secreted or membrane metalloproteinases found in the 1990s. It is the fir
st astacin enzyme found in lymphohematopoietic tissues (kidney, head kidney
, and spleen in fish). Carp nephrosin in its secreted form contains 198
amino acid residues whose predicted relativemolecular weight is 23k. Four puta
tive heparin binding motifs can be identified by which secreted nephrosin ma
y be localized to the heparan sulphate proteoglycan present in extracellular m
atrix. In this thesis, we examined the metal ion requ
irement for the activation of purified nephrosin (apoenzyme). It is concluded
that in addition to the catalytic Zn2+ binding site (Zn site), nephrosi
n contains a Ca2+ binding site since Ca2+ (0.2-0.8mM) can further enhan
ce the activity of nephrosin when it is activated by 100mM Zn2+. Pontential
agonists for the Zn site of nephrosin include Zn2+, Mn2+, Co2+, Cu2+, Ni2+ an
d Cd2+, and potential agonists for the Ca site include Mn2+, Co2+, Cu2+,
Ca2+, Sr2+ and Mg2+. It was also observed that hep
arin reduced nephrosin activity when nephrosin was activated by Zn2+ (100mM
) but lost its inhibitory activity when nephrosin was co-activated by
Zn2+ (100mM) and Ca2+ (500mM). It is postulated that nephrosin is stored in
its mature form but its activity is prohibited by binding to heparin or hepa
rin-like glycosaminoglycan. Once nephrosin is secreted into the extracellula
r fluid, exposure to higher concentrations of extracellular Ca2+ may remo
ve the inhibitory action of heparin and thus convert the inactive form into f
ully active form.
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