Send Orders of Reprints at reprints@benthamscience.net Current Organic Chemistry, 2013, 17, 701-718 701 Enzymatic Synthesis of Oligosaccharides: A Powerful Tool for a Sweet Challenge Marco Filice,* a Marzia Marciello b a Instituto de Catálisis y Petroleoquímica, (CSIC). Campus UAM Cantoblanco, 28049 Madrid, Spain b Instituto de Ciencia de Materiales, (CSIC). Campus UAM Cantoblanco, 28049 Madrid, Spain Abstract: Carbohydrates are complex and structurally diverse compounds in nature with key roles in a broad range of life processes in- cluding signal transduction, carcinogenesis and immune responses. Many natural products contain oligosaccharides that are vital for their biological activity. Despite ongoing challenges, tremendous progresses have been made in recent years for the synthesis of carbohydrates. The chemical glycosylation methods have become more sophisticated and the synthesis of oligosaccharides has become more predict- able. Nonetheless, thanks to their straightforward stereoselectivity and efficiency, carbohydrate-processing enzymes follow being a pow- erful practical alternative in a wide set of synthetic applications targeted to the obtainment of natural oligosaccharides, glycoconjugates and their analogues. In this review, recurring to many practical examples, a general updated overview of the behavior, the advantages and the drawbacks related to the application of glycosyltransferases, glycosylhydrolases and glycosynthases in the oligosaccharide synthesis will be provided. Keywords: Carbohydrate, glycosyltransferase, glycosylhydrolases, glycosynthases, biocatalysis, glycoconjugate. INTRODUCTION Oligosaccharides (or glycans) are a major class of naturally oc- curring carbohydrates, generally, consisting of 3 to 10 monosaccha- rides. Beyond their traditionally accepted roles as energy sources and structural polymers, it is now well established that carbohy- drates, one of the three major classes of biopolymers (together with proteins and nucleic acids), play a pivotal role in numerous biologi- cal processes [1]. In fact, carbohydrates abound on all cell surfaces and in ex- tracellular matrices, as free polysaccharides and as glycoconjugates (i.e., glycan structures directly linked to small molecules, lipids or proteins). Mostly, when glycans are co- or post-translationally ap- pended to the majority of human proteins -process known as glyco- sylation- they form a very important bioconjugate named glycopro- teins. By this way, the structure and function of the proteome is dramatically increased [2]. Analyzing more carefully these chal- lenging biomolecules, it is possible to note how the functional gly- coprotein population is composed by different glycoforms introduc- ing the key concept of glycan heterogeneity. By this term it is pos- sible to describe the evidence that different glycans with significant structural variability decorate an invariant protein sequence. Conse- quently, it has become clearer that the carbohydrate moieties can tune activities and govern specific tasks, both, by modulating the properties of the protein to which they are attached (intrinsically) and by modulating interactions with other biomolecules (extrinsi- cally). And the explanation about how they are able to do so with remarkable control exactly relies on their structural diversity [1c]. Hence, considering the paradoxical small human genome [3], gly- can diversity may explains the human complexity, especially thanks to the structural divergence of glycans being the latter the key to the evolution and speciation of the organisms [3,4]. *Address correspondence to this author at the Instituto de Catálisis y Petroleoquímica, (CSIC). Campus UAM Cantoblanco, 28049 Madrid, Spain; Tel: +34915854870; Fax: +34915854760; E-mail: marcof@icp.csic.es In the eukaryotic organisms, the structural diversity of oligo- saccharides is determined by their biosynthesis in the endoplasmic reticulum (ER) and Golgi apparatus in the cell, in an assembly-line like process that, unlike the protein synthesis, is not template driven and is subject to multiple sequential enzymatic pathways [4a,5]. In such organisms, the bioactive oligosaccharides can commonly be found on the cell surface where they are involved in cell growth and development, cancer cell metastasis, anticoagulation, immune rec- ognition and response, cell–cell communication and initiation of microbial pathogenesis [6]. The biological role of these oligosac- charides relies on their interactions with proteins and on the modu- lation of protein activity at the cell–extracellular interface. Hence, bioactive oligosaccharides have consequently been identified as a medicinally relevant class of biomolecules for the development of therapeutic agents based on specific oligosaccharide structures or mimics thereof [7]. Even in prokaryotic organisms (archaea and bacteria), despite the lack of the same cellular machinery found in eukaryotes, similar processes for oligosaccharide biosynthesis have been conserved. In this case, the bacterial periplasm is equivalent to the eukaryotic ER during biosynthesis of N-linked oligosaccharides whereas the O- linked oligosaccharides are formed in the bacterial cytoplasm or at the interface between cytoplasm and surface appendages such as pili and flagella [8]. These filamentous glycoproteic protrusions of the bacterial surface [9] are crucial elements for the intercellular connection and transfer of plasmids between bacteria (pili) and for the bacterial motility (flagellum). Commonly, the pathogen-mediated infection cascade is initi- ated by the recognition and the binding to the surface of the host cell mediated by the glycoconjugates expressed on the surface of the invading organism. Additionally, many bacteria also release protein toxins that similarly exploit adhesion to cell-surface carbo- hydrates mechanism to penetrate in the target cells. An example of such strategy is represented by the selective binding of cholera toxin from Vibrio cholerae to the cell surface glycolipid ganglioside GM1a (Scheme 1) [10]. 1875-5348/13 $58.00+.00 © 2013 Bentham Science Publishers