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Advances in Glycobiology (virtual)

MED 225/CHEM 237/CMM 225
Offered every two years, Next Offering Spring 2021, First Day March 30, 2021
Location: Virtual meeting via Zoom
Co-Directors: Jeffrey Esko and Kamil Godula
Additional Instructors: Philip Gordts, Jim Paulson, Kevin Campbell
Units: 4
Grading is based on regular attendance and class participation.

*UCSD students and postdocs go to Canvas for assigned reading, study questions, and presentation materials.


Tracy Gilstrap
Education Coordinator
(858) 882-1378

*if you are off campus and would like access to the course materials please contact Tracy
Advanced elective for upper level undergraduates, graduate, and health sciences students who have taken Introduction to Glycoscience (Chem 142/242) and courses in cell biology or biochemistry. This course consists of  discussions of classic papers leading to modern concepts in glycobiology, with the objective of exploring the structure, metabolism, and function of glycans in biological systems. Students should develop a sense of the history of the field, seminal discoveries, and how these discoveries led to changes in the way scientists think about glycans. Students will be divided up into teams and make 1-2 presentations in consultation with a faculty member.

This year the course will be divided up into five modules:

Advances Intro Lecture.pptx
No Class / Attend SDGS

​Module 1: Glycoengineering


​Part 1 - discovery of glycosylation mutants


Regulatory Mutations in CHO Cells Induce Expression of the Mouse Embryonic Antigen SSEA-1.pdf

Isolation of Wheat Germ Agglutinin-resistant Clones of Chinese Hamster Ovary Cells Deficient in Membrane Sialic Acid and Galactose
Part 2 - engineering with glycan remodeling enzymes
Required Reading:
Paper 1: ​Biosynthesis of Mammalian Glycoproteins.pdf
Paper 2: Solubilized glycosyltransferases and biosynthesis in vitro of glycolipids.pdf
Paper 3: Ex vivo glycan engineering of CD44 programs human multipotent mesenchymal stromal cell trafficking to bone.pdf

Recommended Reading:
Paper 4: Expeditious Chemoenzymatic Synthesis of Homogeneous N-Glycoproteins Carrying Defined Oligosaccharide Ligands .pdf
Paper 5: Emerging methods for the production of homogeneous human glycoproteins -review.pdf
Paper 6: Bacterial glycosyltransferase-mediated cell-surface chemoenzymatic glycan modification - recent advances.pdf
Part 3 - metabolic glycan engineering
Required Reading:
​Paper 1: Biosynthesis of a Nonphysiological Sialic Acid in Different Rat Printed inU.S.A. Organs, Using N-Propanoyl-D-hexosaminesas Precursors
Paper 2: Engineering Chemical Reactivity on Cell Surfaces Through Oligosaccharide Biosynthesis.pdf

Recommended Reading:
Paper 3: Metabolic incorporation of unnatural sialic acids into Haemophilus ducreyi lipooligosaccharides.pdf
Paper 4: Versatile Biosynthetic Engineering of Sialic Acid in Living Cells Using Synthetic Sialic Acid Analogues.pdf
Paper 5: Visualizing enveloping layer glycans during zebrafish early embryogenesis.pdf
Paper 6: Metabolic oligosaccharide engineering as a tool for glycobiology - review.pdf
​Part 4 - de novo glycocalyx construction
Required Reading:
Paper 1: Investigations on Cellular Blood-Group Substances.pdf

Paper 2: Noncovalent Cell Surface Engineering/ Incorporation of Bioactive Synthetic Glycopolymers into Cellular Membranes

Recommended Reading: (Please take a look at these papers to be able to discuss the key concepts)
Paper 3: The cancer glycocalyx mechanically primes integrin-mediated growth and survival.pdf

Paper 4: The cancer glycocalyx mechanically primes integrin-mediated growth and survival.pdf

​​Module 2: Flu and Glycan-Protein Interactions

Provided as background material for entire Module:

Adaptation of Influenza Viruses to Human Airway Receptors
A J Thompson & J C Paulson
J Biol Chem 2021;  296:100017. Nov 3;296:100017. doi: 10.1074/jbc.REV120.013309.

Influenza Virus Neuraminidase Structure and Functions
J L McAuley, B P Gilbertson, S Trifkovic, L E Brown, J L McKimm-Breschkin
Front Microbiol 2019 Jan 29;10:39. doi: 10.3389/fmicb.2019.00039. eCollection 2019.

Part 1 - Influenza Virus Receptor Specificity
Required Reading:

The genetic character of O-D change in influenza A
F M Burnet, J D Stone, A Isaacs, M Edney
Br J Exp Pathol 1949 Oct;30(5):419-25.

Receptor specificity in human, avian, and equine H2 and H3 influenza virus isolates
R J Connor, Y Kawaoka, R G Webster, J C Paulson
Virology 1994 Nov 15;205(1):17-23. doi: 10.1006/viro.1994.1615.

Part 2 - Influenza HA and NA balance
​Required Reading:

Amino acid residues contributing to the substrate specificity of the influenza A virus neuraminidase
D Kobasa, S Kodihalli, M Luo, M R Castrucci, I Donatelli, Y Suzuki, T Suzuki, Y Kawaoka
J Virol 1999 Aug;73(8):6743-51. doi: 10.1128/JVI.73.8.6743-6751.1999.

Functional balance of the hemagglutinin and neuraminidase activities accompanies the emergence of the 2009 H1N1 influenza pandemic
R Xu, X Zhu, R McBride, C M Nycholat, W Yu, J C Paulson, I A Wilson
J Virol 2012 Sep;86(17):9221-32. doi: 10.1128/JVI.00697-12. Epub 2012 Jun 20.

​Part 3 - Adaptation of Avian Influenza to human receptors
Required Reading:

Experimental adaptation of an influenza H5 HA confers respiratory droplet transmission to a reassortant H5 HA/H1N1 virus in ferrets
M Imai, T Watanabe, M Hatta, S C Das, M Ozawa, K Shinya, G Zhong, A Hanson, H Katsura, S Watanabe, C Li, E Kawakami, S Yamada, M Kiso, Y Suzuki, E A Maher, G Neumann, Y Kawaoka
Nature 2012 May 2;486(7403):420-8. doi: 10.1038/nature10831.

Three mutations switch H7N9 influenza to
human-type receptor specificity

de Vries RP, Peng W, Grant OC, Thompson AJ, Zhu X, Bouwman KM, de la Pena ATT, van Breemen MJ, Ambepitiya Wickramasinghe IN, de Haan CAM, Yu W, McBride R, Sanders RW, Woods RJ, Verheije MH, Wilson IA, Paulson JC.
PLoS Pathog. 2017 Jun 15;13(6):e1006390. doi: 10.1371/journal.ppat.1006390. eCollection 2017 Jun.
PMID: 28617868 

​Part 4 - Evolution of H3N2 HA receptors and impact on vaccine production

Required Reading:

Recent H3N2 Viruses Have Evolved Specificity for Extended, Branched Human-type Receptors, Conferring Potential for Increased Avidity
W Peng, R P de Vries, O C Grant, A J Thompson, R McBride, B Tsogtbaatar, P S Lee, N Razi, I A Wilson, R J Woods, J C Paulson
Cell Host Microbe 2017 Jan 11;21(1):23-34. doi: 10.1016/j.chom.2016.11.004. Epub 2016 Dec 22.

Contemporary H3N2 influenza viruses have a glycosylation site that alters binding of antibodies elicited by egg-adapted vaccine strains
S J Zost, K Parkhouse, M E Gumina, K Kim, S Diaz Perez, P C Wilson, J J Treanor, A J Sant, S Cobey, S E Hensley
Proc Natl Acad Sci U S A 2017 Nov 21;114(47):12578-12583. doi: 10.1073/pnas.1712377114. Epub 2017 Nov 6.

​Modern Methods in Glycobiology

​General methodological considerations when designing, conducting and interpreting glycoproteomics experiments and examples of recent applications using this technology will be covered.
​Will discuss valuable bioinformatics and systems biology tools relevant to glycobiology

​Module 3: The Heparin / Heparan Sulfate Code

​Provided as background material for entire Module:

ALL students are asked to read the following two reviews on the concept of the heparan sulfate code in preparation for your presentation. 

Deciphering functional glycosaminoglycan motifs in development. (Links to an external site.)
Townley RA, Bülow HE.
Curr Opin Struct Biol. 2018 Jun;50:144-154.

Specificity of glycosaminoglycan-protein interactions. (Links to an external site.)
Kjellén L, Lindahl U.
Curr Opin Struct Biol. 2018 Jun;50:101-108.

​Heparin: Native notions on chemistry and anti-coagulation
Required Reading:
Paper #1: Identification of iduronic acid as the major sulfated uronic acid of heparin
Lindahl U, Axelsson OJ (1971) 
J Biol Chem 246(1):74–82

Paper #2: Anticoagulant action of heparin
Damus PS, Hicks M, Rosenberg RD (1973) Nature 246(5432):355–357

Recommended Reading: 
A Century of Heparin 2019.pdf
Ong CS, Marcum JA, Zehr KJ, Cameron DE.
Ann Thorac Surg. 2019 Sep;108(3):955-958.

220 MHz spectra of heparin, chondroitins, and other mucopolysaccharides 1970.pdf
A. S. Perlin, B. Casu, G. R. Sanderson, and L. F. JohnsonAUTHORS INFO & AFFILIATIONS
Canadian Journal of Chemistry • July 1970 •

The purification and mechanism of action of human antithrombin-heparin cofactor.pdf
Rosenberg RD, Damus PS.
J Biol Chem. 1973 Sep 25;248(18):6490-505.
​Heparin and AT binding demands a sizable structure
Required Reading:

Paper 1:
Evidence for a 3-O-sulfated D-glucosamine residue in the antithrombin-binding sequence of heparin.
Lindahl U, Bäckström G, Thunberg L, Leder IG. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6551-5

Paper 2Anticoagulant activities of heparin oligosaccharides and their neutralization by platelet factor 4
Lane DA, Denton J, Flynn AM, Thunberg L, Lindahl U. Biochem J. 1984 Mar 15;218(3):725-32.

Recommended Reading: 
Anticoagulant activity of heparin: separation of high-activity and low-activity heparin species by affinity chromatography on immobilized antithrombin
Höök M, Björk I, Hopwood J, Lindahl U.
FEBS Lett. 1976 Jul 1;66(1):90-3.2.

Structure of the antithrombin-binding site in heparin
Lindahl U, Bäckström G, Höök M, Thunberg L, Fransson LA, Linker A.
Proc Natl Acad Sci U S A. 1979 Jul;76(7):3198-202.

Role of ternary complexes, in which heparin binds both antithrombin and proteinase, in the acceleration of the reactions between antithrombin and thrombin or factor Xa.
Danielsson A, Raub E, Lindahl U, Björk I.
J Biol Chem. 1986 Nov 25;261(33):15467-73.
​Premise for the heparan sulfate code hypothesis
Required Reading:

Paper 1:
Expression of heparan sulfate D-glucosaminyl 3-O-sulfotransferase isoforms reveals novel substrate specificities.
Liu J, Shworak NW, Sinaÿ P, Schwartz JJ, Zhang L, Fritze LM, Rosenberg RD.
J Biol Chem. 1999 Feb 19;274(8):5185-92.

Paper 2: A novel role for 3-O-sulfated heparan sulfate in herpes simplex virus 1 entry.
Shukla D, Liu J, Blaiklock P, Shworak NW, Bai X, Esko JD, Cohen GH, Eisenberg RJ, Rosenberg RD, Spear PG.
Cell. 1999 Oct 1;99(1):13-22. 

Paper 3Activating and inhibitory heparin sequences for FGF-2 (basic FGF). Distinct requirements for FGF-1, FGF-2, and FGF-4.
Guimond S, Maccarana M, Olwin BB, Lindahl U, Rapraeger AC. J Biol Chem. 1993 Nov 15;268(32):23906-14.

Recommended Reading: 
6-O-sulfotransferase-1 represents a critical enzyme in the anticoagulant heparan sulfate biosynthetic pathway.
Zhang L, Beeler DL, Lawrence R, Lech M, Liu J, Davis JC, Shriver Z, Sasisekharan R, Rosenberg RD. J Biol Chem. 2001 Nov 9;276(45):42311-21.

Cell surface, heparin-like molecules are required for binding of basic fibroblast growth factor to its high affinity receptor.
Yayon A, Klagsbrun M, Esko JD, Leder P, Ornitz DM. Cell. 1991 Feb 22;64(4):841-8.

Crystal structure of fibroblast growth factor receptor ectodomain bound to ligand and heparin.
Pellegrini L, Burke DF, von Delft F, Mulloy B, Blundell TL. Nature. 2000 Oct 26;407(6807):1029-34. 
​Current and future approaches to crack the HS code
Required Reading/Papers to Present:
Paper 1: ZNF263 is a transcriptional regulator of heparin and heparan sulfate biosynthesis.
Weiss RJ, Spahn PN, Toledo AG, Chiang AWT, Kellman BP, Li J, Benner C, Glass CK, Gordts PLSM, Lewis NE, Esko JD.
Proc Natl Acad Sci U S A. 2020 Apr 28;117(17):9311-9317. 

Paper 2: The 3-O-sulfation of heparan sulfate modulates protein binding and lyase degradation.
Chopra P, Joshi A, Wu J, Lu W, Yadavalli T, Wolfert MA, Shukla D, Zaia J, Boons GJ.
Proc Natl Acad Sci U S A. 2021 Jan 19;118(3):e2012935118.

Recommended Reading: 
Genome-wide screens uncover KDM2B as a modifier of protein binding to heparan sulfate.
Weiss RJ, Spahn PN, Chiang AWT, Liu Q, Li J, Hamill KM, Rother S, Clausen TM, Hoeksema MA, Timm BM, Godula K, Glass CK, Tor Y, Gordts PLSM, Lewis NE, Esko JD.
Nat Chem Biol. 2021 Apr 12.

Sequencing Heparan Sulfate Using HILIC LC-NETD-MS/MS.
Wu J, Wei J, Chopra P, Boons GJ, Lin C, Zaia J.
Anal Chem. 2019 Sep 17;91(18):11738-11746.

​​Module 4: Detective Work: Discovery and Elucidation of Dystroglycanopathies

Provided as background material for entire Module:

ALL students are asked to read the following two reviews.

Three Muscular Dystrophies: Review Loss of Cytoskeleton-Extracellular Matrix Linkage

Recent advancements in understanding mammalian O-mannosylation

​Discovery of dystroglycanopathies as a form of muscular dystrophy
Required Reading/Papers to Present:

Ibraghimov-Beskrovnaya O, Ervasti JM, Leveille CJ, Slaughter CA, Sernett SW, Campbell KP. 1992. Primary structure of dystrophin-associated glycoproteins linking dystrophin to the extracellular matrix. Nature, 355:696-702.
DOI: 10.1038/355696a0

Kobayashi K, Nakahori Y, Miyake M, Matsumura K, Kondo-Iida E, Nomura Y, Segawa M, Yoshioka M, Saito K, Osawa M, et al. 1998. An ancient retrotransposal insertion causes Fukuyama-type congenital muscular dystrophy. Nature, 394:388-392. DOI: 10.1038/28653

Michele DE, Barresi R, Kanagawa M, Saito F, Cohn RD, Satz JS, Dollar J, Nishino I, Kelley RI, Somer H, et al. 2002. Post-translational disruption of dystroglycan-ligand interactions in congenital muscular dystrophies. Nature, 418:417-422. DOI: 10.1038/nature00837

Background Reading:
Ervasti JM, Campbell KP. 1993. A role for the dystrophin-glycoprotein complex as a transmembrane linker between laminin and actin. J Cell Biol, 122:809-823.
DOI: 10.1083/jcb.122.4.809

​Genetic analysis of dystroglycanopathies
Required Reading/Papers to Present:

Beltran-Valero de Bernabe D, Currier S, Steinbrecher A, Celli J, van Beusekom E, van der Zwaag B, Kayserili H, Merlini L, Chitayat D, Dobyns WB, et al. 2002. Mutations in the O-mannosyltransferase gene POMT1 give rise to the severe neuronal migration disorder Walker-Warburg syndrome. Am J Hum Genet, 71:1033-1043. DOI: 10.1086/342975

Brockington M, Blake DJ, Prandini P, Brown SC, Torelli S, Benson MA, Ponting CP, Estournet B, Romero NB, Mercuri E, et al. 2001. Mutations in the fukutin-related protein gene (FKRP) cause a form of congenital muscular dystrophy with secondary laminin alpha2 deficiency and abnormal glycosylation of alpha-dystroglycan. Am J Hum Genet, 69:1198-1209.
DOI: 10.1086/324412

Willer T, Lee H, Lommel M, Yoshida-Moriguchi T, de Bernabe DB, Venzke D, Cirak S, Schachter H, Vajsar J, Voit T, et al. 2012. ISPD loss-of-function mutations disrupt dystroglycan O-mannosylation and cause Walker-Warburg syndrome. Nat Genet, 44:575-580.
DOI: 10.1038/ng.2252

Background Reading:

Yoshida A, Kobayashi K, Manya H, Taniguchi K, Kano H, Mizuno M, Inazu T, Mitsuhashi H, Takahashi S, Takeuchi M, et al. 2001. Muscular dystrophy and neuronal migration disorder caused by mutations in a glycosyltransferase, POMGnT1. Developmental Cell, 1:717-724.
DOI: 10.1016/s1534-5807(01)00070-3

Hara Y, Balci-Hayta B, Yoshida-Moriguchi T, Kanagawa M, Beltran-Valero de Bernabe D, Gundesli H, Willer T, Satz JS, Crawford RW, Burden SJ, et al. 2011. A dystroglycan mutation associated with limb-girdle muscular dystrophy. N Engl J Med, 364:939-946.
DOI: 10.1056/NEJMoa1006939

​Elucidation of matriglycan structure and assembly
Required Reading/Papers to Present:

Kanagawa M, Saito F, Kunz S, Yoshida-Moriguchi T, Barresi R, Kobayashi YM, Muschler J, Dumanski JP, Michele DE, Oldstone MB, et al. 2004. Molecular recognition by LARGE is essential for expression of functional dystroglycan. Cell, 117:953-964.
DOI: 10.1016/j.cell.2004.06.003

Inamori K, Yoshida-Moriguchi T, Hara Y, Anderson ME, Yu L, Campbell KP. 2012. Dystroglycan function requires xylosyl- and glucuronyltransferase activities of LARGE. Science, 335:93-96.
DOI: 10.1126/science.1214115

Briggs, D., Yoshida-Moriguchi, T., Zheng, T., Venzke, D., Anderson, M., Strazzulli, A., Moracci, M., Yu, L., Hohenester, E., and Campbell, K.P. Structural Basis of Laminin Binding to the LARGE Glycans on Dystroglycan. Nat. Chem. Biol. 12:810-814, 2016.
DOI: 10.1038/nchembio.2146

Background Reading:

Longman C, Brockington M, Torelli S, Jimenez-Mallebrera C, Kennedy C, Khalil N, Feng L, Saran RK, Voit T, Merlini L, et al. 2003. Mutations in the human LARGE gene cause MDC1D, a novel form of congenital muscular dystrophy with severe mental retardation and abnormal glycosylation of alpha-dystroglycan. Hum Mol Genet, 12:2853-2861.
DOI: 10.1093/hmg/ddg307
Hara Y, Kanagawa M, Kunz S, Yoshida-Moriguchi T, Satz JS, Kobayashi YM, Zhu Z, Burden SJ, Oldstone MB, Campbell KP. 2011. Like-acetylglucosaminyltransferase (LARGE)-dependent modification of dystroglycan at Thr-317/319 is required for laminin binding and arenavirus infection. P Natl Acad Sci USA, 108:17426-17431.
DOI: 10.1073/pnas.1114836108

Goddeeris MM, Wu B, Venzke D, Yoshida-Moriguchi T, Saito F, Matsumura K, Moore SA, Campbell KP. 2013. LARGE glycans on dystroglycan function as a tunable matrix scaffold to prevent dystrophy. Nature, 503:136-140. DOI: 10.1038/nature12605

​Matriglycan function in the central nervous system and neuromuscular junction
Required Reading/Papers to Present:

Moore SA, Saito F, Chen JG, Michele DE, Henry MD, Messing A, Cohn RD, Ross-Barta SE, Westra S, Williamson RA, et al. 2002. Deletion of brain dystroglycan recapitulates aspects of congenital muscular dystrophy. Nature, 418:422-425. DOI: 10.1038/nature00838

Campanelli, J.T., Roberds, S.L., Campbell, K.P. and Scheller, R.H. A Role for Dystrophin-Associated Glycoproteins and Utrophin in Agrin-Induced AChR Clustering. Cell 77:663-674, 1994. DOI: 10.1016/0092-8674(94)90051-5

Grady, R., Zhou, H., Cunningham, J. M., Henry, M. D., Campbell, K. P., & Sanes, J. R. (2000). Maturation and Maintenance of the Neuromuscular Synapse. Neuron, 25(2), 279–293. DOI: 10.1016/s0896-6273(00)80894-6

Background Reading:

Longman C, Brockington M, Torelli S, Jimenez-Mallebrera C, Kennedy C, Khalil N, Feng L, Saran RK, Voit T, Merlini L, et al. 2003. Mutations in the human LARGE gene cause MDC1D, a novel form of congenital muscular dystrophy with severe mental retardation and abnormal glycosylation of alpha-dystroglycan. Hum Mol Genet, 12:2853-2861.
DOI: 10.1093/hmg/ddg307

Saito F, Moore SA, Barresi R, Henry MD, Messing A, Ross-Barta SE, Cohn RD, Williamson RA, Sluka KA, Sherman DL, et al. 2003. Unique role of dystroglycan in peripheral nerve myelination, nodal structure, and sodium channel stabilization. Neuron, 38:747-758.
DOI: 10.1016/s0896-6273(03)00301-5
Ohlendieck, K., Ervasti, J.M., Matsumura, K., Kahl, S.D., Leveille, C.J. and Campbell, K.P. Dystrophin-Related Protein is Localized to Neuromuscular Junctions of Adult Skeletal Muscle. Neuron 7:499-508, 1991.
DOI: 10.1016/0896-6273(91)90301-f

Satz JS, Barresi R, Durbeej M, Willer T, Turner A, Moore SA, Campbell KP. 2008. Brain and eye malformations resembling Walker-Warburg syndrome are recapitulated in mice by dystroglycan deletion in the epiblast. J Neurosci, 28:10567-10575.
DOI: 10.1523/JNEUROSCI.2457-08.2008