Research review paper Carbohydrate synthesis and biosynthesis technologies for cracking of the glycan code: Recent advances Hynek Mrázek, Lenka Weignerová, Pavla Bojarová, Petr Novák, Ondřej Vaněk, Karel Bezouška ⁎ Department of Biochemistry, Faculty of Science, Charles University Prague, Hlavova 8, CZ-12840 Praha 2, Czech Republic Institute of Microbiology, Academy of Sciences of Czech Republic, Vídeňská 1083, CZ-14220 Praha 4, Czech Republic abstract article info Available online xxxx Keywords: Cellular factories Complex carbohydrates Glycodrugs Glycoforms Glycoprotein hormones Protein glycosylation Structure activity studies Vaccination The glycan code of glycoproteins can be conceptually defined at molecular level by the sequence of well char- acterized glycans attached to evolutionarily predetermined amino acids along the polypeptide chain. Func- tional consequences of protein glycosylation are numerous, and include a hierarchy of properties from general physicochemical characteristics such as solubility, stability and protection of the polypeptide from the environment up to specific glycan interactions. Definition of the glycan code for glycoproteins has been so far hampered by the lack of chemically defined glycoprotein glycoforms that proved to be extremely difficult to purify from natural sources, and the total chemical synthesis of which has been hitherto possible only for very small molecular species. This review summarizes the recent progress in chemical and chemoen- zymatic synthesis of complex glycans and their protein conjugates. Progress in our understanding of the ways in which a particular glycoprotein glycoform gives rise to a unique set of functional properties is now having far reaching implications for the biotechnology of important glycodrugs such as therapeutical monoclonal antibodies, glycoprotein hormones, carbohydrate conjugates used for vaccination and other practically im- portant protein–carbohydrate conjugates. © 2012 Elsevier Inc. All rights reserved. Contents 1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 1.1. Defining the glycan code . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 2. Biosynthesis of glycoconjugates with emphasis on glycoproteins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 2.1. Glycoproteins with N-glycosidically linked oligosaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 2.2. Glycoproteins with O-glycosidically linked oligosaccharides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 3. Cellular engineering technologies for the synthesis of defined glycoconjugates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 4. Chemoenzymatic and multienzyme synthesis of complex glycans . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 5. New developments in total chemical synthesis of complex glycan sequences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 6. Progress in the synthesis of carbohydrate conjugates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 6.1. N-Glycosidically linked conjugates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 6.2. O-glycosidically linked conjugates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 7. Analyses of complex oligosaccharides and glycoconjugates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 8. Practical implications of complex glycan bioengineering . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 9. Conclusions and areas for future research . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 0 1. Introduction Protein glycosylation is among the most frequent and abundant pro- tein modifications with numerous functional consequences on protein solubility, stability, folding, assembly into fully active complexes, and Biotechnology Advances xxx (2012) xxx–xxx ⁎ Corresponding author at: Department of Biochemistry, Faculty of Science, Charles University Prague, Hlavova 8, CZ-12840 Praha 2, Czech Republic. Tel.: + 420 2 2195 1273; fax: +420 2 2195 1283. E-mail address: karel.bezouska@natur.cuni.cz (K. Bezouška). JBA-06573; No of Pages 21 0734-9750/$ – see front matter © 2012 Elsevier Inc. All rights reserved. doi:10.1016/j.biotechadv.2012.03.008 Contents lists available at SciVerse ScienceDirect Biotechnology Advances journal homepage: www.elsevier.com/locate/biotechadv Please cite this article as: Mrázek H, et al, Carbohydrate synthesis and biosynthesis technologies for cracking of the glycan code: Recent ad- vances, Biotechnol Adv (2012), doi:10.1016/j.biotechadv.2012.03.008