3I3B

E.coli (lacz) Beta-Galactosidase (M542A) in Complex with D-Galactopyranosyl-1-on

Summary for 3I3B

• Entry DOI: 10.2210/pdb3i3b/pdb
• Related: 1DP0 1JYX 1JZ5 3I3D 3I3E
• Descriptor: Beta-galactosidase, D-galactonolactone, MAGNESIUM ION, ... (6 entities in total)
• Functional Keywords: beta-galactosidase, tim barrel (alpha/beta barrel), jelly-roll barrel, immunoglobulin beta supersandwhich, glycosidase, hydrolase
• Biological source: Escherichia coli
• Total number of polymer chains: 4
• Total formula weight: 476977.14

Authors

Dugdale, M.L.,Dymianiw, D.,Minhas, B.,Huber, R.E. (deposition date: 2009-06-30, release date: 2010-05-12, Last modification date: 2023-09-06)

Primary citation

Dugdale, M.L.,Dymianiw, D.L.,Minhas, B.K.,D'Angelo, I.,Huber, R.E.
Role of Met-542 as a guide for the conformational changes of Phe-601 that occur during the reaction of β-galactosidase (Escherichia coli).
Biochem.Cell Biol., 88:861-869, 2010

PubMed Abstract: The Met-542 residue of β-galactosidase is important for the enzyme's activity because it acts as a guide for the movement of the benzyl side chain of Phe-601 between two stable positions. This movement occurs in concert with an important conformational change (open vs. closed) of an active site loop (residues 794-803). Phe-601 and Arg-599, which interact with each other via the π electrons of Phe-601 and the guanidium cation of Arg-599, move out of their normal positions and become disordered when Met-542 is replaced by an Ala residue because of the loss of the guide. Since the backbone carbonyl of Phe-601 is a ligand for Na(+), the Na(+) also moves out of its normal position and becomes disordered; the Na(+) binds about 120 times more poorly. In turn, two other Na(+) ligands, Asn-604 and Asp-201, become disordered. A substrate analog (IPTG) restored Arg-599, Phe-601, and Na(+) to their normal open-loop positions, whereas a transition state analog d-galactonolactone) restored them to their normal closed-loop positions. These compounds also restored order to Phe-601, Asn-604, Asp-201, and Na(+). Binding energy was, however, necessary to restore structure and order. The K(s) values of oNPG and pNPG and the competitive K(i) values of substrate analogs were 90-250 times higher than with native enzyme, whereas the competitive K(i) values of transition state analogs were ~3.5-10 times higher. Because of this, the E•S energy level is raised more than the E•transition state energy level and less activation energy is needed for galactosylation. The galactosylation rates (k₂) of M542A-β-galactosidase therefore increase. However, the rate of degalactosylation (k₃) decreased because the E•transition state complex is less stable.

PubMed: 20921997
DOI: 10.1139/O10-009

Experimental method

X-RAY DIFFRACTION (2.2 Å)