Structure and mechanism of the chalcogen-detoxifying protein TehB from Escherichia coli.
The oxyanion derivatives of the chalcogens tellurium and selenium are toxic to living organisms even at very low levels. Bacteria have developed mechanisms to overcome their toxicity by methylating them. The structure of TehB from Escherichia coli has been determined in the presence of the cofactor analogues SAH (S-adenosylhomocysteine) and sinefungin (a non-hydrolysable form of S-adenosyl-L-methionine) at 1.48 Å (1 Å=0.1 nm) and 1.9 Å respectively. Interestingly, our kinetic data show that TehB does not discriminate between selenium or tellurite oxyanions, making it a very powerful detoxifying protein. Analysis of the active site has identified three conserved residues that are capable of binding and orientating the metals for nucleophilic attack: His176, Arg177 and Arg184. Mutagenesis studies revealed that the H176A and R184A mutants retained most of their activity, whereas the R177A mutant had 65% of its activity abolished. Based on the structure and kinetic data we propose an SN2 nucleophilic attack reaction mechanism. These data provide the first molecular understanding of the detoxification of chalcogens by bacteria.
Membrane Protein Laboratory, Imperial College London, Diamond Light Source, Chilton, Oxfordshire OX11 0DE, UK.
Hassanul G
HG
Choudhury
Alexander D
AD
Cameron
So
S
Iwata
Konstantinos
K
Beis
D000241:Adenosine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D055162:Biocatalysis; D020134:Catalytic Domain; D018011:Chalcogens / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D016384:Consensus Sequence; D018360:Crystallography, X-Ray; D024881:Drug Resistance, Bacterial; D048168:Escherichia coli K12 / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008745:Methylation; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D016566:Organoselenium Compounds / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012435:S-Adenosylhomocysteine / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013691:Tellurium / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
D018011:Chalcogens; D029968:Escherichia coli Proteins; D050505:Mutant Proteins; D016566:Organoselenium Compounds; D011994:Recombinant Proteins; C119920:tellurite resistance protein B, E coli; D012435:S-Adenosylhomocysteine; D000241:Adenosine; D013691:Tellurium; C006235:sinefungin