Exchange of glutamine-217 to Glutamate of Clostridium Limosum Exoenzyme C3 Turns the Asparagine-Specific ADP-ribosyltransferase Into an Arginine-Modifying Enzyme

Biochemistry. 2006 Jan 24;45(3):1017-25. doi: 10.1021/bi052253g.

Abstract

C3-like ADP-ribosyltransferaseses are produced by Clostridium species, Bacillus cereus, and various Staphylococcus aureus strains. The exoenzymes modify the low-molecular-mass GTPases RhoA, B, and C. In structural studies of C3-like exoenzymes, an ARTT-motif (ADP-ribosylating turn-turn motif) was identified that appears to be involved in substrate specificity and recognition (Han, S., Arvai, A. S., Clancy, S. B., Tainer, J. A. (2001) J. Mol. Biol. 305, 95-107). Exchange of Gln217, which is a key residue of the ARTT-motif, to Glu in C3 from Clostridium limosum results in inhibition of ADP-ribosyltransferase activity toward RhoA. The mutant protein is still capable of NAD-binding and possesses NAD+ glycohydrolase activity. Whereas recombinant wild-type C3 modifies Rho proteins specifically at an asparagine residue (Asn41), Gln217Glu-C3 is capable of ADP-ribosylation of poly-arginine but not poly-asparagine. Soybean trypsin inhibitor, a model substrate for many arginine-specific ADP-ribosyltransferases, is modified by the Gln217Glu-C3 transferase. Also in C3 ADP-ribosyltransferases from Clostridium botulinum and B. cereus, the exchange of the equivalent Gln residue to Glu blocked asparagine modification of RhoA but elicited arginine-specific ADP-ribosylation. Moreover, the Gln217Glu-C3lim transferase was able to ADP-ribosylate recombinant wild-type C3lim at Arg86, resulting in decrease in ADP-ribosyltransferase activity of the wild-type enzyme. The data indicate that the exchange of one amino acid residue in the ARTT-motif turns the asparagine-modifying ADP-ribosyltransferases of the C3 family into arginine-ADP-ribosylating transferases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • ADP Ribose Transferases / chemistry
  • ADP Ribose Transferases / genetics
  • ADP Ribose Transferases / metabolism*
  • Amino Acid Sequence
  • Arginine / metabolism*
  • Asparagine / metabolism*
  • Botulinum Toxins / metabolism*
  • Clostridium / enzymology*
  • Clostridium / genetics
  • Escherichia coli / genetics
  • Glutamic Acid / metabolism*
  • Glutamine / metabolism*
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship
  • Substrate Specificity
  • Trypsin Inhibitor, Bowman-Birk Soybean

Substances

  • Recombinant Proteins
  • Trypsin Inhibitor, Bowman-Birk Soybean
  • Glutamine
  • Glutamic Acid
  • Asparagine
  • Arginine
  • ADP Ribose Transferases
  • exoenzyme C3, Clostridium botulinum
  • Botulinum Toxins