2RKS

Crystal structure of Staphylococcal nuclease variant PHS L38K at cryogenic temperature

Summary for 2RKS

• Entry DOI: 10.2210/pdb2rks/pdb
• Related: 2RBM
• Descriptor: Thermonuclease, PHOSPHATE ION (3 entities in total)
• Functional Keywords: staphylococcal nuclease, hyperstable variant, hydrolase, internal ion pair
• Biological source: Staphylococcus aureus
• Cellular location: Nuclease A: Secreted. Nuclease B: Membrane: P00644
• Total number of polymer chains: 1
• Total formula weight: 16968.24

Authors

Schlessman, J.L.,Harms, M.J.,Garcia-Moreno, E.B. (deposition date: 2007-10-17, release date: 2008-04-15, Last modification date: 2024-04-03)

Primary citation

Harms, M.J.,Schlessman, J.L.,Chimenti, M.S.,Sue, G.R.,Damjanovic, A.,Garcia-Moreno, B.
A buried lysine that titrates with a normal pKa: role of conformational flexibility at the protein-water interface as a determinant of pKa values.
Protein Sci., 17:833-845, 2008

PubMed Abstract: Previously we reported that Lys, Asp, and Glu residues at positions 66 and 92 in staphylococcal nuclease (SNase) titrate with pK(a) values shifted by up to 5 pK(a) units in the direction that promotes the neutral state. In contrast, the internal Lys-38 in SNase titrates with a normal pK(a). The crystal structure of the L38K variant shows that the side chain of Lys-38 is buried. The ionizable moiety is approximately 7 A from solvent and ion paired with Glu-122. This suggests that the pK(a) value of Lys-38 is normal because the energetic penalty for dehydration is offset by a favorable Coulomb interaction. However, the pK(a) of Lys-38 was also normal when Glu-122 was replaced with Gln or with Ala. Continuum electrostatics calculations were unable to reproduce the pK(a) of Lys-38 unless the protein was treated with an artificially high dielectric constant, consistent with structural reorganization being responsible for the normal pK(a) value of Lys-38. This reorganization must be local because circular dichroism and NMR spectroscopy indicate that the L38K protein is native-like under all conditions studied. In molecular dynamics simulations, the ion pair between Lys-38 and Glu-122 is unstable. The simulations show that a minor rearrangement of a loop is sufficient to allow penetration of water to the amino moiety of Lys-38. This illustrates both the important roles of local flexibility and water penetration as determinants of pK(a) values of ionizable groups buried near the protein-water interface, and the challenges faced by structure-based pK(a) calculations in reproducing these effects.

PubMed: 18369193
DOI: 10.1110/ps.073397708

Experimental method

X-RAY DIFFRACTION (2.01 Å)