The P-loop--a common motif in ATP- and GTP-binding proteins.

The crystal structure of the H-ras oncogene protein p21 complexed to the slowly hydrolysing GTP analogue GppNp has been determined at 1.35 A resolution. 211 water molecules have been built into the electron density. The structure has been refined to a final R-factor of 19.8% for all data between 6 A and 1.35 A. The binding sites of the nucleotide and the magnesium ion are revealed in high detail. For the stretch of amino acid residues 61-65, the temperature factors of backbone atoms are four times the average value of 16.1 A2 due to the multiple conformations. In one of these conformations, the side chain of Gln61 makes contact with a water molecule, which is perfectly placed to be the nucleophile attacking the gamma-phosphate of GTP. Based on this observation, we propose a mechanism for GTP hydrolysis involving mainly Gln61 and Glu63 as activating species for in-line attack of water. Nucleophilic displacement is facilitated by hydrogen bonds from residues Thr35, Gly60 and Lys16. A mechanism for rate enhancement by GAP is also proposed.

http://cbm.msoe.edu/includes/pdf/2008RasPrimCit.pdf

Refined crystal structure of the triphosphate conformation of H-ras p21 at 1.35 Å resolution : implications for the mechanism of GTP hydrolysis

Conformational constraints for amyloid fibrillation: the importance of being unfolded.

Publisher Summary This chapter provides an insight that quantitative predictions of the stability and cooperative folding behavior of proteins are within reach. The problem of predicting the structure of a protein from its amino acid sequence constitutes the classical “protein folding problem.” The thermodynamic aspects of this problem constitute the focus of the chapter. Experimental thermodynamic values for the fundamental forces that determine protein stability are refined to a point in which accurate calculations of the free energy of stabilization are possible, provided that the protein structure is known. The chapter also (1) predicts the overall stability of proteins based on structural information, (2) predicts the cooperative folding/unfolding behavior of proteins, (3) predicts the magnitude of the interactions among structural domains in proteins, and (4) predicts the presence of specific partially folded intermediates. Estimation of the folding/unfolding partition function using a hierarchical approximation and subsequent prediction of cooperative behavior is also increasingly feasible, as demonstrated for myoglobin and yeast phosphoglycerate kinase. Thermodynamic stability and cooperative interactions in proteins can be determined by hierarchical approach to partition functions, thermodynamic dissection of cooperative interactions, and classification of cooperative forces and mechanisms. The strategy used to dissect specific fundamental interactions is to isolate first the hydrophobic effects and subsequently other interactions. Some future work will extend the capability to proteins exhibiting other than pure α-helical structural motifs.

Thermodynamics of structural stability and cooperative folding behavior in proteins.

The role of the α-helix dipole in protein function and structure

The structure of yeast phosphoglycerate kinase has been determined with data obtained from amino acid sequence, nucleotide sequence, and X-ray crystallographic studies. The substrate binding sites, as deduced from electron density maps, are compatible with known substrate specificity and the stereochemical requirements for the enzymic reaction. A carboxyl-imidazole interaction appears to be involved in controlling the transition between the open and closed forms of the enzyme.

P. L. Wendell

Papers

Sequence and structure of yeast phosphoglycerate kinase.

Sequence andstructure ofyeastphosphoglycerate kinase