Cloning, purification and characterization of Geobacillus stearothermophilus V uroporphyrinogen-III C-methyltransferase: evaluation of its role in resistance to potassium tellurite in Escherichia coli.
The Geobacillus stearothermophilus V cobA gene encoding uroporphyrinogen-III C-methyltransferase (also referred to as SUMT) was cloned into Escherichia coli and the recombinant enzyme was overexpressed and purified to homogeneity. The enzyme binds S-adenosyl-L-methionine and catalyzes the production of III methyl uroporphyrinogen in vitro. E. coli cells expressing the G. stearothermophilus V cobA gene exhibited increased resistance to potassium tellurite and potassium tellurate. Site-directed mutagenesis of cobA abolished tellurite resistance of the mesophilic, heterologous host and SUMT activity in vitro. No methylated, volatile derivatives of tellurium were found in the headspace of tellurite-exposed cobA-expressing E. coli, suggesting that the role of SUMT methyltransferase in tellurite(ate) detoxification is not related to tellurium volatilization.
Araya
Manuel A
MA
Facultad de Química y Biología, Universidad de Santiago de Chile, Chile. manuel.araya@roche.com
Tantaleán
Juan C
JC
Pérez
José M
JM
Fuentes
Derie E
DE
Calderón
Iván L
IL
Saavedra
Claudia P
CP
Burra
Radhika
R
Chasteen
Thomas G
TG
Vásquez
Claudio C
CC
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism*; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D008780:Methyltransferases / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000096:biosynthesis; D012436:S-Adenosylmethionine / Q000378:metabolism; D013691:Tellurium / Q000378:metabolism*; D014577:Uroporphyrinogens / Q000096:biosynthesis
D011994:Recombinant Proteins; D014577:Uroporphyrinogens; C026899:potassium tellurate(IV); D012436:S-Adenosylmethionine; D008780:Methyltransferases; C060508:uroporphyrin-III C-methyltransferase; D013691:Tellurium