Post-translational formylglycine modification of bacterial sulfatases by the radical S-adenosylmethionine protein AtsB.
C(alpha)-Formylglycine (FGly) is the catalytic residue of sulfatases. FGly is generated by post-translational modification of a cysteine (prokaryotes and eukaryotes) or serine (prokaryotes) located in a conserved (C/S)XPXR motif. AtsB of Klebsiella pneumoniae is directly involved in FGly generation from serine. AtsB is predicted to belong to the newly discovered radical S-adenosylmethionine (SAM) superfamily. By in vivo and in vitro studies we show that SAM is the critical co-factor for formation of a functional AtsB.SAM.sulfatase complex and for FGly formation by AtsB. The SAM-binding site of AtsB involves (83)GGE(85) and possibly also a juxtaposed FeS center coordinated by Cys(39) and Cys(42), as indicated by alanine scanning mutagenesis. Mutation of these and other conserved cysteines as well as treatment with metal chelators fully impaired FGly formation, indicating that all three predicted FeS centers are crucial for AtsB function. It is concluded that AtsB oxidizes serine to FGly by a radical mechanism that is initiated through reductive cleavage of SAM, thereby generating the highly oxidizing deoxyadenosyl radical, which abstracts a hydrogen from the serine-C(beta)H(2)-OH side chain.
Fang
Qinghua
Q
Institut für Biochemie und Molekulare Zellbiologie, Abteilung Biochemie II, Universität Göttingen, Heinrich-Düker-Weg 12, 37073 Göttingen, Germany.
Peng
Jianhe
J
Dierks
Thomas
T
D000409:Alanine / Q000737:chemistry; D020816:Amino Acid Motifs; D001419:Bacteria / Q000201:enzymology*; D001665:Binding Sites; D002614:Chelating Agents / Q000494:pharmacology; D003545:Cysteine / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006859:Hydrogen / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000378:metabolism / Q000502:physiology*; D007711:Klebsiella pneumoniae / Q000378:metabolism; D008956:Models, Chemical; D016297:Mutagenesis, Site-Directed; D010100:Oxygen / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D012436:S-Adenosylmethionine / Q000737:chemistry*; D013429:Sulfatases / Q000737:chemistry*; D013696:Temperature; D013997:Time Factors
C471554:AtsB protein, Klebsiella pneumoniae; D002614:Chelating Agents; D007506:Iron-Sulfur Proteins; D010455:Peptides; D012436:S-Adenosylmethionine; D006859:Hydrogen; D005982:Glutathione Transferase; D013429:Sulfatases; D003545:Cysteine; D000409:Alanine; D010100:Oxygen