C. Sousa et al., METALLOADSORPTION BY ESCHERICHIA-COLI-CELLS DISPLAYING YEAST AND MAMMALIAN METALLOTHIONEINS ANCHORED TO THE OUTER-MEMBRANE PROTEIN LAMB, Journal of bacteriology, 180(9), 1998, pp. 2280-2284
Yeast (CUP1) and mammalian (HMT-1A) metallothioneins (MTs) have been e
fficiently expressed in Escherichia call as fusions to the outer membr
ane protein LamB. A 65-amino-acid sequence from the CUP1 protein of Sa
ccharomyces cerevisiae (yeast [Y] MT) was genetically inserted in perm
issive site 153 of the LamB sequence, which faces the outer medium. A
second LamB fusion at position 153 was created with 66 amino acids rec
ruited from the form of human (H) MT that is predominant in the adipos
e tissue, HMT-1A. Both LamB(153)-YMT and LamB(153)-HMT hybrids were pr
oduced in vivo as full-length proteins, without any indication of inst
ability or proteolytic degradation. Each of the two fusion proteins wa
s functional as the port of entry of lambda phage variants, suggesting
maintenance of the overall topology of the wild-type LamB. Expression
of the hybrid proteins in vivo multiplied the natural ability. of E.
coli cells to bind Cd2+ 15- to 20-fold, in good correlation with the n
umber of metal-binding centers contributed by the MT moiety of the fus
ions.