Enzymatic synthesis of UDP-(3-deoxy-3-fluoro)-D-galactose and UDP-(2-deoxy-2-fluoro)-D-galactose and substrate activity with UDP-galactopyranose mutase

被引:51
作者
Barlow, JN [1 ]
Blanchard, JS [1 ]
机构
[1] Albert Einstein Coll Med, Dept Biochem, Bronx, NY 10461 USA
关键词
UDP-sugars; fluoro-sugars; tautomerization; UDP-galactopyranose mutase; uridine 5 '-(3-deoxy-3-fluoro-D-galactopyranosyl diphosphate); UDP-(2-deoxy-2-fluoro)-galactose;
D O I
10.1016/S0008-6215(00)00135-X
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The novel UDP-sugar uridine 5'-(3-deoxy-3-fluoro-D-galactopyranosyl diphosphate) (I) and UDP-(2-deoxy-2-fluoro)-D-galactose (2) have been prepared enzymatically and tested as substrate analogues for the enzyme UDP-galactopyranose mutase (UDP-Galp mutase EC 5.4.99.9). Turnover of both 1 and 2 by UDP-Galp mutase was observed by HPLC and F-19 NMR. The HPLC elution profile and F-19 chemical shift of the products are consistent with the formation of the predicted furanose forms of 1 and 2. The K-m values for compounds 1 and 2 were similar to those of the natural substrate UDP-Galp (0.26 mM for 1, 0.2 mM for 2, and 0.6 mM for UDP-Galp), but the values for k(cat) were substantially different (1.6/min for I, 0.02/min for 2, and 1364/min for UDP-Galp). A correlation was also observed between the equilibrium yield of product formed during turnover of UDP-sugar by UDP-Galp mutase (UDP-Galp, compound 1 or compound 2), and the amount of furanose present for the free sugar at thermal equilibrium in aqueous solution, using H-1 and F-19 NMR spectroscopy. The implications of these results to the mechanism of the unusual enzymatic reaction are discussed. (C) 2000 Elsevier Science Ltd. All rights reserved.
引用
收藏
页码:473 / 480
页数:8
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