QM/MM study of β-1,4-galactosyltransferase-1

M. Krupicka¹, I. Tvaroska¹

¹Slovak Academy of Sciences, Institute of Chemistry, Dubravska cesta 9, Bratislava, Slovak Republic

chemmakr@savba.sk

In the absence of α-lactalbumin, the enzyme β-1,4-galactosyltransferase-1 (Gal-T1) catalyses transfer of galactose residue from UDP-Gal to the C-4 hydroxyl group of N-acetlyglucosamine. The catalytic mechanism of Gal-T1 was investigated using hybrid quantum mechanical/molecular mechanical (QM/MM) method with QM part containing 253 atoms treated with density functional theory (DFT) at the BP/DZP level. The remaining parts of Gal-T1 complex, altogether 4527 atoms were modeled using the AMBER molecular force field. A theoretical model of Michaelis complex was built using the X-ray structures of Gal-T1 containing donor or acceptor substrate respectively. The QM(DFT)/MM model identified a S_N2-type transition state with D318 as the catalytic base for the reaction in the enzyme active site. In the transition state (TS), donor sugar is almost fully cleaved from pyrophosphate, while nucleophilic oxygen O4 remains protonated, with low barrier hydrogen bond transfer to the catalytic base. The structure of TS is characterized by the O_A4-C1 and C1-O1 distances 2.703 Å and 2.092 Å respectively. The activation barrier for the proposed reaction was estimated to be ~14 kcal/mol. This modeling study provides detailed insight into the mechanism of the Gal transfer catalyzed by Gal-T1.

Acknowledgement: This work has been supported by MRTN-CT-2006-035866 (REVCAT) and VEGA 2/0176/09