Rohitesh Gupta1, Khushi L Matta2, Sriram Neelamegham3. 1. Department of Chemical and Biological Engineering, State University of New York, Buffalo, NY 14260, USA. Electronic address: rohitesh.gupta@gmail.com. 2. Tumor End LLC, Baton Rouge, LA 70803, USA. 3. Department of Chemical and Biological Engineering, State University of New York, Buffalo, NY 14260, USA; NY State Center for Excellence in Bioinformatics and Life Sciences, State University of New York, Buffalo, NY 14260, USA. Electronic address: neel@buffalo.edu.
Abstract
Sialyltransferases (STs) catalyze the addition of sialic acids to the non-reducing ends of glycoproteins and glycolipids. In this work, we examined the acceptor specificity of five human α(2,3)sialyltransferases, namely ST3Gal -I, -II, -III, -IV and -VI. KM values for each of these enzymes is presented using radioactivity for acceptors containing Type-I (Galβ1,3GlcNAc), Type-II (Galβ1,4GlcNAc), Type-III (Galβ1,3GalNAc) and Core-2 (Galβ1,3(GlcNAcβ1,6)GalNAc) reactive groups. Several variants of acceptors inhibited ST3Gal activity emphasizing structural role of acceptor in enzyme-catalyzed reactions. In some cases, mass spectrometry was performed for structural verification. The results demonstrate human ST3Gal-I catalysis towards Type-III and Core-2 acceptors with KM = 5-50 μM and high VMax values. The KM for ST3Gal-I and ST3Gal-II was 100 and 30-fold lower, respectively, for Type-III compared to Type-I acceptors. Variants of Type-I and Type-II structures characterized ST3Gal-III, -IV and -VI for their catalytic specificity. This manuscript also estimates KM for human ST3Gal-VI using Type-I and Type-II substrates. Together, these findings built a platform for designing inhibitors of STs having therapeutic potential.
Sialyltransferases (STs) catalyze the addition of n class="Chemical">sialic acids to the non-reducing ends of glycoproteins and glycolipids. In this work, we examined the acceptor specificity of five human α(2,3)sialyltransferases, namely ST3Gal -I, -II, -III, -IV and -VI. KM values for each of these enzymes is presented using radioactivity for acceptors containing Type-I (Galβ1,3GlcNAc), Type-II (Galβ1,4GlcNAc), Type-III (Galβ1,3GalNAc) and Core-2 (Galβ1,3(GlcNAcβ1,6)GalNAc) reactive groups. Several variants of acceptors inhibited ST3Gal activity emphasizing structural role of acceptor in enzyme-catalyzed reactions. In some cases, mass spectrometry was performed for structural verification. The results demonstrate humanST3Gal-I catalysis towards Type-III and Core-2 acceptors with KM = 5-50 μM and high VMax values. The KM for ST3Gal-I and ST3Gal-II was 100 and 30-fold lower, respectively, for Type-III compared to Type-I acceptors. Variants of Type-I and Type-II structures characterized ST3Gal-III, -IV and -VI for their catalytic specificity. This manuscript also estimates KM for humanST3Gal-VI using Type-I and Type-II substrates. Together, these findings built a platform for designing inhibitors of STs having therapeutic potential.
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