1D4F

CRYSTAL STRUCTURE OF RECOMBINANT RAT-LIVER D244E MUTANT S-ADENOSYLHOMOCYSTEINE HYDROLASE

Summary for 1D4F

• Entry DOI: 10.2210/pdb1d4f/pdb
• Related: 1B3R
• Descriptor: S-ADENOSYLHOMOCYSTEINE HYDROLASE, NICOTINAMIDE-ADENINE-DINUCLEOTIDE, ADENOSINE, ... (4 entities in total)
• Functional Keywords: s-adenosylhomocysteine hydrolase, adohcyase, adohcy, mutagenesis, x-ray crystal structure, enzyme structure, hydrolase
• Biological source: Rattus norvegicus (Norway rat)
• Cellular location: Cytoplasm: P10760
• Total number of polymer chains: 4
• Total formula weight: 193641.62

Authors

Komoto, J.,Huang, Y.,Takusagawa, F.,Gomi, T.,Ogawa, H.,Takata, Y.,Fujioka, M. (deposition date: 2000-06-22, release date: 2001-01-17, Last modification date: 2024-02-07)

Primary citation

Komoto, J.,Huang, Y.,Gomi, T.,Ogawa, H.,Takata, Y.,Fujioka, M.,Takusagawa, F.
Effects of site-directed mutagenesis on structure and function of recombinant rat liver S-adenosylhomocysteine hydrolase. Crystal structure of D244E mutant enzyme.
J.Biol.Chem., 275:32147-32156, 2000

PubMed Abstract: A site-directed mutagenesis, D244E, of S-adenosylhomocysteine hydrolase (AdoHcyase) changes drastically the nature of the protein, especially the NAD(+) binding affinity. The mutant enzyme contained NADH rather than NAD(+) (Gomi, T., Takata, Y., Date, T., Fujioka, M., Aksamit, R. R., Backlund, P. S., and Cantoni, G. L. (1990) J. Biol. Chem. 265, 16102-16107). In contrast to the site-directed mutagenesis study, the crystal structures of human and rat AdoHcyase recently determined have shown that the carboxyl group of Asp-244 points in a direction opposite to the bound NAD molecule and does not participate in any hydrogen bonds with the NAD molecule. To explain the discrepancy between the mutagenesis study and the x-ray studies, we have determined the crystal structure of the recombinant rat-liver D244E mutant enzyme to 2.8-A resolution. The D244E mutation changes the enzyme structure from the open to the closed conformation by means of a approximately 17 degrees rotation of the individual catalytic domains around the molecular hinge sections. The D244E mutation shifts the catalytic reaction from a reversible to an irreversible fashion. The large affinity difference between NAD(+) and NADH is mainly due to the enzyme conformation, but not to the binding-site geometry; an NAD(+) in the open conformation is readily released from the enzyme, whereas an NADH in the closed conformation is trapped and cannot leave the enzyme. A catalytic mechanism of AdoHcyase has been proposed on the basis of the crystal structures of the wild-type and D244E enzymes.

PubMed: 10913437
DOI: 10.1074/jbc.M003725200

Experimental method

X-RAY DIFFRACTION (2.8 Å)