Expression of a Neurospora crassa metallothionein and its variants in Escherichia coli.
Appl Environ Microbiol
; 56(9): 2748-54, 1990 Sep.
Article
en En
| MEDLINE
| ID: mdl-2148862
ABSTRACT
The Neurospora crassa metallothionein (NC) synthesis gene was cloned and expressed in Escherichia coli in two different expression vectors (pING2 and pUA7), both under the regulation of the Salmonella typhimurium arabinose operon. Upon induction with arabinose, the pING2-NC vector expressed as inclusion body-localized AraB'NC fusion protein of 21 kilodaltons. The pUA7-NC vector expressed a 5.3-kilodalton LppNC fusion protein anchored to the outer membrane of the cell. Cells expressing the NC fusion proteins accumulated Cd2+ and Cu+ (between 2.3- and 11-fold) compared with nonexpressing cells. To generate novel forms of metal-binding peptides, a set of specific mutant genes for N. crassa NC was designed in which each cysteine residue was replaced with a subset of amino acids implicated in peptide-metal coordination (Asn, Asp, His, Lys, or Tyr residues). These mutant NC sequences were cloned into the two vectors and expressed in E. coli. One of the mutant proteins (containing His residues) showed accumulation of Cd2+ and Cu+ (threefold) from a mixture of 16 heavy metals species. None of the other heavy metals present in the culture was accumulated.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Metalotioneína
/
Neurospora crassa
Idioma:
En
Revista:
Appl Environ Microbiol
Año:
1990
Tipo del documento:
Article