METALLOADSORPTION BY ESCHERICHIA-COLI-CELLS DISPLAYING YEAST AND MAMMALIAN METALLOTHIONEINS ANCHORED TO THE OUTER-MEMBRANE PROTEIN LAMB

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.