Structure of the regulatory subunit of CK2 in the presence of a p21WAF1 peptide demonstrates flexibility of the acidic loop.

A truncated form of the regulatory subunit of the protein kinase CK2beta (residues 1-178) has been crystallized in the presence of a fragment of the cyclin-dependent kinase inhibitor p21WAF1 (residues 46-65) and the structure solved at 2.9 A resolution by molecular replacement. The core of the CK2beta dimer shows a high structural similarity with that identified in previous structural analyses of the dimer and the holoenzyme. However, the electron density corresponding to the substrate-binding acidic loop (residues 55-64) indicates two conformations that differ from that of the holoenzyme structure [Niefind et al. (2001), EMBO J. 20, 5320-5331]. Difference electron density near the dimerization region in each of the eight protomers in the asymmetric unit is attributed to between one and eight amino-acid residues of a complexed fragment of p21WAF1. This binding site corresponds to the solvent-accessible part of the conserved zinc-finger motif.

Structure of E. coli ketopantoate hydroxymethyl transferase complexed with ketopantoate and Mg2+, solved by locating 160 selenomethionine sites.

We report the crystal structure of E. coli ketopantoate hydroxymethyltransferase (KPHMT) at 1.9 A resolution, in complex with its product, ketopantoate. KPHMT catalyzes the first step in the biosynthesis of pantothenate (vitamin B(5)), the precursor of coenzyme A and the acyl carrier protein cofactor. The structure of the decameric enzyme was solved by multiwavelength anomalous dispersion to locate 160 selenomethionine sites and phase 560 kDa of protein, making it the largest structure solved by this approach. KPHMT adopts the (betaalpha)(8) barrel fold and is a member of the phosphoenolpyruvate/pyruvate superfamily. The active site contains a ketopantoate bidentately coordinated to Mg(2+). Similar binding is likely for the substrate, alpha-ketoisovalerate, orienting the C3 for deprotonation.

Cockroach allergen Bla g 2: structure, function, and implications for allergic sensitization.

Exposure to German cockroach (Blattella germanica) allergens is associated with the development of chronic respiratory diseases, especially asthma. The mechanism by which allergic patients develop specific immunoglobulin E (IgE) responses to environmental allergens is unknown. However, recent reports provided evidence that enzyme activity, especially proteolytic activity, was a major contributor to allergenicity. Bla g 2 is one of the most potent cockroach allergens (prevalence of IgE responses of 60 to 80%) and shows homology to the aspartic proteinase family of enzymes. We investigated whether the allergenicity of Bla g 2 was linked to its putative enzymatic function. A molecular model of Bla g 2, based on the high resolution crystal structures of pepsin and chymosin, showed that the overall three-dimensional structure of Bla g 2 was similar to that of aspartic proteinases with a well-defined binding pocket. However, critical amino acid substitutions in the catalytic triads and in the "flap" region of the molecule suggested that Bla g 2 was inactive and homologous to mammalian pregnancy-associated glycoproteins. This was confirmed experimentally by enzyme assay. The results show dissociation between enzymatic activity and allergenicity for Bla g 2 and suggest that other genetic and environmental factors are important determinants of sensitization.