1UIP

ADENOSINE DEAMINASE (HIS 238 GLU MUTANT)

Summary for 1UIP

• Entry DOI: 10.2210/pdb1uip/pdb
• Descriptor: ADENOSINE DEAMINASE, ZINC ION, PURINE RIBOSIDE, ... (4 entities in total)
• Functional Keywords: hydrolase, nucleotide metabolism, zinc cofactor tim barrel
• Biological source: Mus musculus (house mouse)
• Cellular location: Cell membrane; Peripheral membrane protein; Extracellular side (By similarity): P03958
• Total number of polymer chains: 1
• Total formula weight: 40024.80

Authors

Wilson, D.K.,Quiocho, F.A. (deposition date: 1996-08-30, release date: 1997-06-24, Last modification date: 2024-02-14)

Primary citation

Sideraki, V.,Wilson, D.K.,Kurz, L.C.,Quiocho, F.A.,Rudolph, F.B.
Site-directed mutagenesis of histidine 238 in mouse adenosine deaminase: substitution of histidine 238 does not impede hydroxylate formation.
Biochemistry, 35:15019-15028, 1996

PubMed Abstract: His 238, a conserved amino acid located in hydrogen-bonding distance from C-6 of the substrate in the active site of murine adenosine deaminase (mADA) and postulated to play an important role in catalysis, was altered into an alanine, a glutamate, and an arginine using site-directed mutagenesis. The Ala and Glu substitutions did not result in changes of the secondary or tertiary structure, while the Arg mutation caused local perturbations in tertiary structure and quenched the emission of one or more enzyme tryptophans. Neither the Glu or Arg mutations affected substrate binding affinity. By contrast, the Ala mutation enhanced substrate and inhibitor binding by 20-fold. The most inactive of the mutants, Glu 238, had a kcat/K(m) 4 x 10(-6) lower than the wild-type value, suggesting that a positive charge on His 238 is important for proper catalytic function. The Ala 238 mutant was the most active ADA, with a kcat/K(m) 2 x 10(-3) lower than the wild-type value. NMR spectroscopy and crystallography revealed that this mutant is able to catalyze hydration of purine riboside, a ground-state analog of the reaction. These results collectively show that His 238 is not required for formation of the hydroxylate used in the deamination and may instead have an important electrostatic role.

PubMed: 8942668
DOI: 10.1021/bi961427e

Experimental method

X-RAY DIFFRACTION (2.4 Å)