Interactions between the enzyme and its ligands in the active site of β-galactosidase

Andrea Štěpánková^{1, 2}, Tereza Skálová², Jan Dohnálek², Jarmila Dušková², Petr Kolenko^{1, 2}, Jindřich Hašek², Vojtěch Spiwok³, Petra Lipovová³

¹ Dept. of Solid State Physics, FNSPE, CTU, Trojanova 13, 120 00, Prague 2, Czech Republic

² Institute of Macromolecular Chemistry AS CR, v.v.i., Heyrovského nám. 2, 162 06, Prague 6, Czech Republic

³ Dept. of Biochemistry, ICT, Technická 5, 166 28, Prague 6, Czech Republic

Stepanko@imc.cas.cz

β-Galactosidase (EC 3.2.1.23) is an enzyme which is able to catalyze hydrolysis of the terminal β-d-galactosyl of β-d-galactosides and it is also able to catalyze trans-glycosylation and belongs to the enzyme class called glycosidases.

Recently, we determined the structure of the wild type enzyme β-galactosidase from Arthrobacter sp. C2-2¹. Structures of its complexes with β-d-galactose, β-d-galactonolactone and isopropyl β-d-1-thiogalactopyranoside (IPTG) determined by X-ray diffraction are presented here. X-ray diffraction data were collected at the beam-line ID14.1 of the source of synchrotron radiation ESRF in Grenoble and on an in-house rotating anode diffractometer. The data were processed using HKL2000. All the three crystals belong to the same space group P2₁ but packing of hexamers in the crystals differs. The asymetric unit contains one hexamer with a molecular weight of 660 kDa.

This paper reviews the interactions between several type of ligands and the enzyme. Our structures of complexes of the enzyme from Arthrobacter sp. C2-2 are compared with a series of complexes of β-galactosidase from E. coli².

There are two distinct binding modes for the galactosyl group of substrate - shallow and deep. Each binding mode has specific hydrogen bonds between enzyme and bound ligands. Structure changes evoked by binding of ligands into the active site of the enzyme will be shown and discussed. β-d-galactose, as well as β-d-galactonolactone, occupy the deep binding position and make the same interactions but large structure changes with regard to non-liganded enzyme are observed only in the case of β-d-galactonolactone.

1. Skálová, T. et al.: Cold-active β-galactosidase from Arthrobacter sp. C2-2 forms compact 660 kDa hexamers: Crystal structure at 1.9 Å resolution. J. Mol. Biol., 353, 282-294 (2005).

2. Juers D. H. et al.: A structural view of the action of Escherichia coli (lacZ) β-galactosidase, Biochemistry, 40, 14781-14794 (2001).

Acknowledgements:

This work was supported by the Czech Science Foundation (305/07/1073) and by the IGS CTU (CTU0803914).