PMID:14633979
| Citation |
Schmitzberger, F, Kilkenny, ML, Lobley, CM, Webb, ME, Vinkovic, M, Matak-Vinkovic, D, Witty, M, Chirgadze, DY, Smith, AG, Abell, C and Blundell, TL (2003) Structural constraints on protein self-processing in L-aspartate-alpha-decarboxylase. EMBO J. 22:6193-204 |
|---|---|
| Abstract |
Aspartate decarboxylase, which is translated as a pro-protein, undergoes intramolecular self-cleavage at Gly24-Ser25. We have determined the crystal structures of an unprocessed native precursor, in addition to Ala24 insertion, Ala26 insertion and Gly24-->Ser, His11-->Ala, Ser25-->Ala, Ser25-->Cys and Ser25-->Thr mutants. Comparative analyses of the cleavage site reveal specific conformational constraints that govern self-processing and demonstrate that considerable rearrangement must occur. We suggest that Thr57 Ogamma and a water molecule form an 'oxyanion hole' that likely stabilizes the proposed oxyoxazolidine intermediate. Thr57 and this water molecule are probable catalytic residues able to support acid-base catalysis. The conformational freedom in the loop preceding the cleavage site appears to play a determining role in the reaction. The molecular mechanism of self-processing, presented here, emphasizes the importance of stabilization of the oxyoxazolidine intermediate. Comparison of the structural features shows significant similarity to those in other self-processing systems, and suggests that models of the cleavage site of such enzymes based on Ser-->Ala or Ser-->Thr mutants alone may lead to erroneous interpretations of the mechanism. |
| Links |
PubMed PMC291833 Online version:10.1093/emboj/cdg575 |
| Keywords |
Alanine; Amino Acid Substitution; Crystallography, X-Ray; Escherichia coli/enzymology; Escherichia coli Proteins/chemistry; Escherichia coli Proteins/metabolism; Glutamate Decarboxylase/chemistry; Glutamate Decarboxylase/metabolism; Models, Molecular; Mutagenesis, Insertional; Mutagenesis, Site-Directed; Protein Processing, Post-Translational; Protein Structure, Secondary; Recombinant Proteins/chemistry; Recombinant Proteins/metabolism |
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