S. Goenka et Cm. Rao, Expression of recombinant zeta-crystallin in Escherichia coli with the help of GroEL/ES and its purification, PROT EX PUR, 21(2), 2001, pp. 260-267
z-Crystallin is a taxon-specific crystallin found in the eye lens of guinea
pig and other hystricomorph rodents and camelids. It is an NADPH:quinone o
xidoreductase and is also present in low amounts in other tissues where it
might act as a detoxifying enzyme. A lens-specific promoter confers lens-sp
ecific expression of the gene in high amounts where it is speculated to pla
y a structural role in maintaining the transparency of the lens ensemble. A
deletion mutation leads to autosomal dominant congenital cataract and also
results in the loss of NADPH binding. In order to perform structural studi
es with the protein with an aim to delineate the cause of cataract in these
mutant guinea pigs, recombinant zeta -crystallin was cloned and expressed
in Escherichia coli. The overexpression of the protein in E. coli resulted
in a major fraction of it partitioning into inclusion bodies. The co-overex
pression of the bacterial chaperone system GroEL/ES along with zeta -crysta
llin could significantly enhance the yield of soluble protein. Active zeta
-crystallin could then be purified from the E. coli using Mono Q anion exch
ange FPLC and was found to be identical to the native zeta -crystallin isol
ated from the guinea pig lens with respect to size, spectral properties, an
d activity. (C) 2001 Academic Press