2GAR

A PH-DEPENDENT STABLIZATION OF AN ACTIVE SITE LOOP OBSERVED FROM LOW AND HIGH PH CRYSTAL STRUCTURES OF MUTANT MONOMERIC GLYCINAMIDE RIBONUCLEOTIDE TRANSFORMYLASE

2GAR の概要

• エントリーDOI: 10.2210/pdb2gar/pdb
• 分子名称: GLYCINAMIDE RIBONUCLEOTIDE TRANSFORMYLASE, PHOSPHATE ION (3 entities in total)
• 機能のキーワード: purine biosynthesis, folate cofactors, loop flexibility, monomer-dimer association, enzyme mechanism, anti-cancer agents
• 由来する生物種: Escherichia coli
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 23303.19

構造登録者

Su, Y.,Yamashita, M.M.,Greasley, S.E.,Mullen, C.A.,Shim, J.H.,Jennings, P.A.,Benkovic, S.J.,Wilson, I.A. (登録日: 1998-05-13, 公開日: 1998-08-12, 最終更新日: 2024-05-29)

主引用文献

Su, Y.,Yamashita, M.M.,Greasley, S.E.,Mullen, C.A.,Shim, J.H.,Jennings, P.A.,Benkovic, S.J.,Wilson, I.A.
A pH-dependent stabilization of an active site loop observed from low and high pH crystal structures of mutant monomeric glycinamide ribonucleotide transformylase at 1.8 to 1.9 A.
J.Mol.Biol., 281:485-499, 1998

PubMed Abstract: A mutation in the dimer interface of Escherichia coli glycinamide ribonucleotide transformylase (GarTfase) disrupts the observed pH-dependent association of the wild-type enzyme, but has no observable effect on the enzyme activity. Here, we assess whether a pH effect on the enzyme's conformation is sufficient by itself to explain the pH-dependence of the GarTfase reaction. A pH-dependent conformational change is observed between two high-resolution crystal structures of the Glu70Ala mutant GarTfase at pH 3.5 (1.8 A) and 7.5 (1.9 A). Residues 110 to 131 in GarTfase undergo a transformation from a disordered loop at pH 3.5, where the enzyme is inactive, to an ordered loop-helix structure at pH 7.5, where the enzyme is active. The ordering of this flexible loop-helix has a direct effect on catalytic residues in the active site, binding of the folate cofactor and shielding of the active site from solvent. A main-chain carbonyl oxygen atom from Tyr115 in the ordered loop forms a hydrogen bond with His108, and thereby provides electronic and structural stabilization of this key active site residue. Kinetic data indicate that the pKa of His108 is in fact raised to 9. 2. The loop movement can be correlated with elevation of the His pKa, but with further stabilization, probably from Asp144, after the binding of folate cofactor. Leu118, also in the loop, becomes positioned near the p-amino benzoic acid binding site, providing additional hydrophobic interactions with the cofactor 10-formyl tetrahydrofolate. Thus, the pH-dependence of the enzyme activity appears to arise from local active site rearrangements and not from differences due to monomer-dimer association.

PubMed: 9698564
DOI: 10.1006/jmbi.1998.1931

実験手法

X-RAY DIFFRACTION (1.8 Å)