Extended Sugar-Assisted Glycopeptide Ligations:  Development, Scope, and Applications (original) (raw)

Toward fully synthetic, homogeneous glycoproteins: Advances in chemical ligation

Biopolymers, 2010

Traditionally, in the pharma sciences, there has been an unstated but operative bifurcation into small molecules and biologics. Small molecules were seen to be, at the discovery level, in the province of chemistry, based on targets provided through biology. By contrast, "biologics" were seen to arise solely from the province of biology exploiting its accessible replicative mechanisms. Our laboratory has been dedicated to the proposition that explosive advances in chemical synthesis have been such as to render so called "biologics" as being accessible to chemical synthesis. In this paper, we focus particularly on the area of glycopeptides. Chemical synthesis, in principle, offers an advantage, in that it can lead to homogeneous glycopeptides characterized by a single glycoform of the glycosidic domain mounted at a particular amino acid in the polypeptide domain. In support of this defining goal, a variety of new methods have been developed. The key problem addressed is that of ligation. In this paper, we review how insights available from mechanistic organic chemistry have been used to create an imposing framework for the synthesis of structures which would, in an earlier day, have been seen to be strictly in the realm of chemically inaccessible "biologics". We dedicate this paper to the remarkable accomplishments of Professor Steven Kent-particularly, his discovery of NCL, which changed the course of peptide ligations.

Sugar-Assisted Glycopeptide Ligation with Complex Oligosaccharides: Scope and Limitations

Journal of the American Chemical Society, 2008

We have previously shown sugar-assisted ligation (SAL) to be a useful method for the convergent construction of glycopeptides. However to date SAL has only been carried out on systems where the thiol auxiliary is attached to a monosaccharide. For SAL to be truly applicable to the construction of fully elaborated glycopeptides and glycoproteins it must be possible to carry out the reaction when the thiol auxiliary is attached to more elaborate sugars, as these are frequently what are observed in nature. Here we examine the effects of glycosylation at C-3, C-4 and C-6 of the C-2 auxiliarycontaining glycan. Model glycopeptides where synthesized chemoenzymatically and reacted with peptide thioesters used in our previous work. These studies reveal that SAL is sensitive to extended glycosylation on the auxiliary-containing sugar. While it is possible to carry out SAL with extended glycosylation at C-4 and C-6, the presence of glycosylation at C-3 prevents the ligation from occurring. Additionally, with glycosylation at C-4 the ligation efficiency is affected by the identity of the N-terminal AA, while the nature of the C-terminal residue of the peptide thioester does not appear affect ligation efficiency. These studies provide useful guidelines in deciding when it is appropriate to use SAL in the synthesis of complex glycopeptides and glycoproteins, and how to design ligation junctions for optimal yield.

Chemical and Chemoenzymatic Synthesis of Glycoproteins for Deciphering Functions

Chemistry & Biology, 2014

This study describes the synthesis of the a-and b-linked N-acetyllactosamine (Galp-b-1,4-GlcNAc; LacNAc) glycosides of threonine (LacNAc-Thr). LacNAc-a-Thr was prepared by direct chemical coupling of a 2-azido-2-deoxy-lactose disaccharide donor to a suitable partially protected threonine unit. In contrast, stepwise chemical generation of b-linked N-acetylglucosamine followed by enzymatic galactosylation to give LacNAc-b-Thr proved effective, whereas use of a 2-azido-2-deoxy-lactose donor in acetonitrile failed to give the desired b-linked disaccharyl glycoside. This study illustrates that it is possible to overcome the inherent stereoselection for 1,2-trans chemical glycosylation with a GlcNAc donor, and that the well-established preference of bovine b-1,4-galactosyltransferase for b-linked acceptor substrates can also be overcome. Using this knowledge, short glycopeptide fragments based on T. cruzi mucin sequences, Thr-Thr-[LacNAcThr]-Thr-Thr-Gly, were synthesised. All LacNAc-based compounds outlined were shown to serve as acceptor substrates for sialylation by T. cruzi trans-sialidase. † The a-glycoside appears to be formed directly, not through equilibration of the b-glycoside. Addition of pure b-glycoside to the reaction mixture did not result in additional a-glycoside formation. straightforward and direct approach to the a-linked GlcNAc-Thr system that, given that there is no need to manipulate C-2 functionality, is competitive with other approaches to such a-linked 2-amino-2-deoxy-glucosides. 30,31 Removal of the benzyl ester and Fmoc groups from protected glyco-amino acids (9) and (10) was performed by standard hydrogenation (10% Pd-C/H 2 ); subsequent removal of the acetate groups was realised in the presence of 1 M NaOMe in MeOH. In this way, 20-30 mg quantities of the completely deprotected glyco-amino acids (3) were obtained (Scheme 1).

Glycopeptide/Glycoprotein Synthesis

Protein glycosylation has dramatically enriched both the functional and structural diversity of proteins in mammals. Due to the non-template biosynthesis of glycoproteins in the nature, glyco-proteins always exist as heterogeneous mixtures with different glycan structures, which have complicated the isolation of pure and well-defined glycoforms for detailed mechanistic and functional studies. Over the past years, different strategies including chemical and chemoenzymatic methods have been developed for obtaining the homogenous glycoproteins, which will offer new opportunities to conduct an extensive assessment of relationship between the structure and function of glycoproteins and to reveal the biological role of the individual oligosaccharide on a glycoprotein for the evolution and development of cells. This chapter gives insight into the recent progress in the development of chemical and chemoenzymatic synthesis of homogenous glycoproteins including native chemical ligation, serine/threonine ligation, and chemoenzymatic glycoprotein remodeling. Examples are selected to demonstrate successful applications of synthetic strategies developed so far.

Convergent Synthesis of N-Linked Glycopeptides on a Solid Support

Journal of the American Chemical Society, 1998

An efficient N-linked glycosylation reaction between glycosylamines and p-nitrophenyl thioester peptides has been developed. The reaction conditions are mild and compatible with the C-terminal free carboxylic acid group and the unprotected N-linked sialyloligosaccharide. By means of this convergent strategy, a versatile N-glycopeptide fragment containing an Nterminal Thz and a C-terminal thioester was readily prepared, which is available for the synthesis of long glycopeptides and glycoproteins using the protocol of native chemical ligation.