Rational design, synthesis, and characterization of novel inhibitors for human beta1,4-galactosyltransferase.
An affinity labeling reagent, uridine 5'-(6-amino-{2-[(7-bromomethyl-2-naphthyl)methoxycarbonylmethoxy]ethoxy}acetyl-6-deoxy-alpha-D-galactopyranosyl) diphosphate (1a), was designed on the basis of 3D docking simulation and synthesized to investigate the functional role of Trp310 residue located in the small loop near the active site of human recombinant galactosyltransferase (betaGalT-1). Mass spectrometric analysis revealed that the Trp310 residue of betaGalT1 can be selectively modified with the naphthylmethyl group of compound 1a at the C-3 position of the indole ring. This result motivated us to synthesize novel uridine-5'-diphosphogalactose (UDP-Gal) analogues as candidates for mechanism-based inhibitors for betaGalT-1. We found that uridine 5'-(6-O-[10-(2-naphthyl)-3,6,9-trioxadecanyl]-alpha-d-galactopyranosyl) diphosphate (2) is the strongest inhibitor (K(i) = 1.86 microM) against UDP-Gal (Km = 4.91 microM) among compounds reported previously. A cold spray ionization time-of-flight mass spectrometry study demonstrated that the complex of this inhibitor and betaGalT-1 cannot interact with an acceptor substrate in the presence of Mn2+.
Takaya
Kenji
K
Division of Biological Sciences, Frontier Research Center for Post-Genome Science and Technology, Graduate School of Science, Hokkaido University, N-21, W-11, Sapporo 001-0021, Japan.
Nagahori
Noriko
N
Kurogochi
Masaki
M
Furuike
Tetsuya
T
Miura
Nobuaki
N
Monde
Kenji
K
Lee
Yuan Chuan
YC
Nishimura
Shin-Ichiro
S
D000345:Affinity Labels / Q000138:chemical synthesis* / Q000737:chemistry; D001665:Binding Sites; D015195:Drug Design; D005700:Galactosyltransferases / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D006801:Humans; D008958:Models, Molecular; D015394:Molecular Structure; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014364:Tryptophan / Q000737:chemistry; D014539:Uridine Diphosphate Sugars / Q000138:chemical synthesis* / Q000737:chemistry
D000345:Affinity Labels; D011994:Recombinant Proteins; D014539:Uridine Diphosphate Sugars; C505170:uridine 5'-(6-O-(10-(2-naphthyl)-3,6,9-trioxadecanyl)galactopyranosyl) diphosphate; C505169:uridine 5'-(6-amino-(2-((7-bromomethyl-2-naphthyl)methoxycarbonylmethoxy)ethoxy)acetyl-6-deoxygalactopyranosyl) diphosphate; D014364:Tryptophan; D005700:Galactosyltransferases; C493310:beta1,4-galactosyltransferase, human