Binding Studies of α-GalNAc-specific Lectins to the α-GalNAc (Tn-antigen) Form of Porcine Submaxillary Mucin and Its Smaller Fragments

Abstract

Isothermal titration microcalorimetry (ITC) and hemagglutination inhibition measurements demonstrate that a chemically and enzymatically prepared form of porcine submaxillary mucin that possesses a molecular mass of ϳ10 6 daltons and ϳ2300 ␣-GalNAc residues (Tn-PSM) binds to the soybean agglutinin (SBA) with a K d of 0.2 nM, which is ϳ10 6 -fold enhanced affinity relative to GalNAc␣1-O-Ser (Tn), the pancarcinoma carbohydrate antigen. The enzymatically derived 81 amino acid tandem repeat domain of Tn-PSM containing ϳ23 ␣-GalNAc residues binds with ϳ10 3 -fold enhanced affinity, while the enzymatically derived 38/40 amino acid cleavage product(s) of Tn-PSM containing ϳ11-12 ␣-GalNAc residues shows ϳ10 2 -fold enhanced affinity. A natural carbohydrate decorated form of PSM (Fd-PSM) containing 40% of the core 1 blood group type A tetrasaccharide, and 58% peptide-linked GalNAc␣1-O-Ser/Thr residues, with 45% of the peptide-linked ␣-GalNAc residues linked ␣- Recently, isothermal titration microcalorimetry (ITC) has been used to investigate the binding of asialofetuin (ASF), a globular glycoprotein that possesses nine LacNAc epitopes on three N-linked triantennary chains, to a number of galectins including galectin-1 (5). The observed K a values for ASF binding to the galectins are 50 -80-fold greater than that of monovalent LacNAc. These results are similar to the increases in * This work was supported by Grants CA-16054 (to C. F. B.), , and Core Grant P30 CA-13330 from the NCI, National Institutes of Health, Dept. of Health, Education, and Welfare. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Evaluation of Riproximin Binding Properties Reveals a Novel Mechanism for Cellular Targeting

“…For aBSM, a molecular mass of 400 kDa was assumed (25,26). 920 GalNAc residues/molecule were estimated for aBSM based on data available for porcine submaxillary mucin, which possesses a molecular mass of 10 6 Da with ϳ2300 GalNAc residues/molecule (20).…”

“…When the interactions of riproximin with each of the proteins ASF and aBSM is considered, the main features of the mechanism of lectins binding to multivalent targets apply (19,20). On multivalent structures, several equivalent epitopes are available for the so-called bind-and-jump effects; the lectin can be recaptured by each of the remaining free epitopes, leading to decreased dissociation.…”

Density-dependent Lectin–Glycan Interactions as a Paradigm for Conditional Regulation by Posttranslational Modifications

“…6 -fold enhanced affinity relative to monovalent GalNAc (19). The high affinity of the mucin is due to the large number of its glycans.…”

A formal approach to the molecular fuzzy lock-and-key

“…While proteins are constructed from 20 basic amino acids, the glycan kelp bed is formed from as many as 7,000 glycan determinants, and represents a vastly more complex system for information transmission (Cummings, 2009;Gupta et al, 2010). Figure 3, from Dam et al (2007), illustrates a 'bind-andslide' mechanism by which increasing concentration of a lectin species can induce a phase transition topological change. Initially, the lectin diffuses along and off the glycan kelp frond, until a sufficient number of sites are occupied.…”

“…Levels of sialome complexity, from core and core modifications to the shifting, bending, twisting, glycan 'kelp fronds' that coat most cell surfaces and, via lectin interaction, constitute sophisticated logic gates involved in explosively vast information transfers: in comparison with the 20 amino acids making up all proteins, some 7,000 glycan determinants are needed to constitute the flexible kelp fronds, side branches and all (Cummings, 2009). Dam et al (2007). Lectin diffuses along and off the flexible glycan frond, until a sufficient number of sites are occupied.…”