2017-2018 CREST Teams
Goucher College (Baltimore MD)
Hello! We are the Goucher College BCMB Club's CREST team in Baltimore, Maryland. Our members consist of Dr. Judy Levine, Kelly Budge, Shannara Bauer, Brandon Creed, Olivia Dickert, and Edna Ferreira. We are all seniors here and began our school's chapter last fall. We are very excited to join the modeling project this year. We plan on modeling OGA interacting with the inhibitor Thiamet-G which has been shown to reduce symptoms of Alzheimer's Disease in a mouse model.
O-GlcNAcase (OGA) with Thiamet-G
PDB file: 5un9
Primary Citation: B. Li, H. Li, L. Lu, J. Jiang, Structures of human O-GlcNAcase and its complexes reveal a new substrate recognition mode. Nature Structural & Molecular Biology. 24, 362 (2017).
Abstract: Alzheimer’s Disease is associated with the hyperphosphorylation of the microtubule-associated protein tau. Protein phosphorylation events often occur at the same positions on a protein as O-GlcNac modifications. The mutually exclusive events exist in a dynamic equilibrium that is regulated by enzyme activity. O-GlcNAcase (OGA) is an enzyme which removes O-GlcNacylation from proteins. Thiamet-G was designed to cross the blood brain barrier as a selective inhibitor of OGA in order to effectively decrease the removal of O-GlcNAcylation events. By inhibiting the ability of OGA, there is evidence that Thiamet-G can lead to a decrease in hyperphosphorylation of tau in vertebrate brains and potentially serve as an effective treatment of Alzheimer’s Disease. We modeled the inhibition of OGA by Thiamet-G to highlight the enzyme residues that interact with the inhibitor and the parts of the protein that are important for recognizing target proteins.
Lane College (Jackson TN)
Greetings. Lane College Science Club joined the ASBMB Student Chapters last year. Our club participates in a few projects, including outreach at Lincoln Elementary School. In this photo, we just finished a program with fourth graders demonstrating Euglena and phototaxis. We are excited to be working on the CREST project building a model of human O-GlcNAcase. Our team members are Le'Juan Berry (Senior), Ashley Gayle (Senior), Darrius Hawkins (Senior), Ryan Billings (Freshman), Ashlee Martin-Richardson (Freshman), and Ihouma Tasie (Senior). Our faculty advisors are Dr. Melanie Van Stry and Dr. Candace Jones.
Human O-GlcNAc Hydrolase (hOGA)
PDB File: 5m7s
Primary Citation. Roth C, Chan S, Offen WA, Hemsworth GR, Willems LI, King DT, Varghese V, Britton R, Vocadlo DJ, Davies GJ. Structural and functional insight into human O-GlcNAcase. Nat Chem Biol. 2017 June; 13(6): 610–612.
Abstract: The post-translational modification (PTM) of proteins enable cells to react promptly to internal and external signals through direct and progressive control of protein function. Addition of O-linked-n-acetylglucosamine (O-GlcNAc) to serine and threonine residues of cytoplasmic, nuclear and mitochondrial proteins is a nourishment and stress response PTM. O-GlcNAcylation has been linked to several diseases such as diabetes and cancer. For example, increased O-GlcNAcylation is directly linked to insulin resistance and to hyperglycemia-induced glucose toxicity, two characteristics of diabetes and diabetic complications. O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA) are enzymes that control the dynamic cycling of this PTM. Here, we have constructed a 3D model of human OGA bound to the transition state analog Thiamet-G using Jmol and the PDB file 5M7S, a truncated version of the enzyme called Split1 (Roth et al. Nat. Chem. Biol. 2017). Split1 forms a functional homodimer that is stabilized by a helical bundle formed by helices from each subunit. Thiamet-G fits in the substrate binding pocket formed by Cys215, Tyr298, and Trp278 and interacts with Gly67, Lys98, Asn280, Glu295, and Asn313 to form hydrogen bonds. Thiamet-G also interacts with the catalytic residues Glu174 and Glu175. The focus of this project was to explore the protein structure of OGA and design and build a physical model that illustrated key functional features of the protein. Funded in part by NSF-DUE 1725940 for the CREST Project.
Nova Southeastern University (Fort Lauderdale FL)
Hello Everyone! We are a group of three students working with our faculty member, Dr. Emily Schmitt Lavin. We just became a chapter of ASBMB this fall with a small group of students. We are looking forward to working on the O-GlcNAcylation modeling project with everyone on the CREST Conversation team. Our group consists of three pre-med students: Sophia Nguyen and Vivian Perez Hernandez who are seniors, and Alesa Chabbra who is a freshman.
OGT in complex with UDP and fused substrate peptide TAB
PDB file: 5lvv
Primary Citation: Rafie K, Raimi O, Ferenbach AT, Borodkin VS, Kapuria V, van Aalten DMF. 2017 Recognition of a glycosylation substrate by the O-GlcNAc transferase TPR repeats. Open Biol. 7:170078. http://dx.doi.org/10.1098/rsob.170078
Abstract: O-GlcNAcylation is a post-translational modification similar in importance to the mechanism of phosphorylation in its ability to affect signal transduction. This process is mediated by the enzyme, O-GlcNAc transferase (OGT). OGT catalyzes the addition of the sugar, N-acetylglucosamine (GlcNAc) from the carrier molecule uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) to certain serine or threonine residues in more than a thousand target substrate proteins. The TAB1 (transforming growth factor-beta-activated kinase 1 binding protein) substrate was fused to OGT. GlcNAc was shown binding to three serine residues in TAB1. Six tetratricopeptide (TPR) repeats were identified. These alpha helical paired repeats fold together to produce a single domain called the TPR domain near the N terminus of OGT. Within the TPR domain, five Asparagine (Asp) residues were identified that are involved in holding the substrate in place. Beta sheets in OGT were also indicated. If TAB1 does not receive GlcNAc it will not be able to signal the proper response of the innate immune system.
Saint Leo University (St. Leo FL)
Greetings from Saint Leo! Our team consists of our faculty member Dr. Audrey Shor and six students. Saint Leo has been a chapter of ASBMB since 2011, however this year's student group is completely new. It includes Corey Gallen, a senior, Allison Cobb, a junior, and Anapaula Rios-Rosales, Isabella Jacus, Raheim Grant and Zarrar Rashman, all first year students. Saint Leo's team is interested in modeling the O-GlcNAcase protein. We look forward to working on this project and with all other teams!
O-GlcNAc Hydrolase (OGA) with bound transition state analog ThiametG
PDB file: 5m7s
Primary Citation: Roth C, Chan S, Offen WA, et al. Structural and functional insight into human O-GlcNAcase. Nat Chem Biol. 2017;13(6):610-612. doi:10.1038/nchembio.2358.
Abstract: A basic component of learning undergraduate biology concepts is grasping the significant interplay between carbohydrates and proteins. O-GlcNacylation is a post-translation modification that covalently attaches O-linked N-acetylglocosamine (O-GlcNAc) moieties to cellular proteins. O-GlcNAc is a sugar moiety that can posttransationally modify serine and threonine residues of target proteins. Much can be gained from studying the targets of O-GlcNacylation. Two enzymes regulate O-GlcNacylation; O-GlcNAc transferase (OGT), which catalyzes the addition of GlcNAc to specific serine and threonine residues, and O-GlcNacylase (OGA), which catalyzes the hydrolysis of the monosaccharide from the substrate. The protein we emphasize, OGA, is a homodimer, with each subunit comprised of 916 residues. Two isoforms of the enzyme exist in humans; 1 and 3 which are expressed in the cytoplasm and nucleus, respectively and are speculated to have different sensitivities to substrate based on their environment. The cytoplasmic isoform includes a full length protein (OGA-L), while the nuclear isoform is shorter (OGA-S), lacks a C-terminal acetyltransferase-like domain and demonstrates deduced enzymatic activity in vitro. Human OGA possesses a fugitive substrate binding groove that is highly conserved among metazoan OGAs, in addition to a stalk domain near the active site. Residues significant to O-GlcNAc, a hydrolase, are close to the sugar/catalytic machinery or occupy positions shown to undergo a conformational change upon inhibitor binding. Mutations targeting the substrate binding groove identified residues that are not required for hydrolysis but serve roles in recognition and binding of substrate. The differences between the residues required for the turnover of these substrates may reflect a difference in molecular movement in OGA sites. Further elucidation of this mechanism could help advance our treatment and/or prevention of diseases like neurological disorders and diabetes.
University of Minnesota Rochester (Rochester MN)
We have been working on our project since early in the 2017 fall semester. We are excited about our CREST project. Our team is of 8 students who have a love and curiosity in biochemistry and medicine! Here's a picture of our tiny university in downtown Rochester since I don't have a group picture.
PDB file: 4gz5
Primary Citation: Lazarus, M., Jiang, J., Gloster, T., Zandberg, W., Whitwort, G., Vocadio, D., & Walker, S. (2012). Structural snapshots of the reaction coordinate for O-GlcNAc transferase. Nat. Chem. Biol., 8, 966-68. Doi: 10.1038/nchembio.1109
Abstract: Currently in the United States more than five million individuals live with Alzheimer’s disease (AD), which can lead to memory loss, difficulty with problem solving, a decreased quality of life and eventually death. AD is initiated by the formation of amyloid plaques in the extracellular space of the brain, however, the reason for this is unknown. The amyloid-ꞵ precursor protein (APP), an integral membrane protein concentrated in neural synapses, has been linked to the development of these plaques. In healthy individuals, the cleavage of APP leads to a long, secreted form of APP (sAPPα) and C-terminal fragments (CTFs). In individuals with AD, cleavage of APP leads to sAPPꞵ, different CTFs, and amyloid-beta (Aꞵ) fragments. The misfolding of Aꞵ contributes to the development of AD when these Aꞵ peptides aggregate and form amyloid plaques. Previous research demonstrated that addition of O-linked N-acetylglucosamine (OGlcNAc) to Thr576 on APP decreases the number of Aꞵ peptides formed, thus O-GlcNAcylation of this protein could be useful for therapeutics and warrants further research. OGlcNAcylation occurs when the enzyme, OGlcNAc Transferase (OGT) (PDB ID: 4gz5), catalyzes covalent attachment of an N-acetylglucosamine molecule to either a serine or threonine residue via a uridine diphosphate N-acetylglucosamine (UDP-OGlcNAc). The UDP enables the transfer mechanism by acting as a good leaving group. Our model depicts OGT bound to UDP-OGlcNAc via Gln839, His498, and Lys898 that will attach OGlcNAc to Thr576 on a flexible toober that models APP. The APP toober, will demonstrate the cleavage and folding of Aꞵ peptide.
University of San Diego (San Diego CA)
O-GlcNAc Transferase: Exploring the Structural Basis of the Cross-Talk between GlcNAcylation and Phosphorylation Using Physical Models
PDB file: 4gyy
Primary Citation: Lazarus, M.B., et. al, Structural snapshots of the reaction coordinate for OGlcNAc transferase. (2012) Nat. Chem.Biol. 8: 966-968
Abstract: The focus of this project was to explore structure-function relationships in the enzyme UDP-OGlcNAc transferase protein by designing a model that illustrates key functional features of the protein, which could then be physically printed using polychromatic coloring 3D printers. In particular our focus is on the Cross-Talk between O-GlcNAcylation and Phosphorylation that plays a critical role in overall regulation. Using the pdb file 4gyy.pdb from the paper “Crystal structure of human O-GlcNAc Transferase with UDP-5SGlcNAc and a peptide substrate” The structure illustrates the binding sites for both UDP-GlcNAc and the peptide YPGGSTPVSSANMM with the requisite PV prior to the GlcNAcylatable S. Recent work by Leny et al. has shown that the preceding T can be phosphorylated and that the sequence TPVS is common in proteins thought to undergo GlcNAc- Phosphorylation cross talk. We have also used computational models to create and built models of the peptide phosphorylated at the N-3 Threonine. These models clearly illustrate that the phosphate group clashes with the U in UDPGlcNAC. Since the kinetic mechanism of OGT is ordered Bi-Bi with UDPGlcNAc as the obligate first substrate, the phosphorylated peptide can no longer bind in the peptide substrate pocket explaining why excludes GlcNAcylation. The models described here can be used to illustrate three different aspects contained in the ASBMB 8 Core Concepts of Macromolecular Structure and Function: #3. Structure and function are related, #4. Macromolecular interactions, and #6. The biological activity of macromolecules is often regulated. In addition construction and use of the models illustrates the use of core concept #8, A variety of experimental and computational approaches can be used to observe and quantitatively measure the structure, dynamics and function of biological macromolecule. Funded in part by NSF-DUE 1725940 for the CREST Project.
University of Wisconsin - Stevens Point (Stevens Point WI)
We finally took a picture :) Our CREST Team is made up of Noah, Nick, Garrick, Cheemeng, and Elisa. Dr. Lawrence is the faculty advisor for our ASBMB "biochemistry club" chapter and helped to recruit students from his biochemistry class. But Dr. Jonsson is leading the charge for the CREST project. We're most interested in studying how the human O-GlcNAc transferase interacts with so many other proteins.
O-linked N-acetylglucosamine transferase (OGT)
PDB file: 5lwv
Primary Citation: Rafie K, Raimi O, Ferenbach AT, Borodkin VS, Kapuria V, van Aalten DMF. 2017 Recognition of a glycosylation substrate by the O-GlcNAc transferase TPR repeats. Open Biol. 7: 170078.
Abstract: O-linked N-acetylglucosamine transferase (OGT) is an enzyme that transfers N-acetylglucosamine (GlcNAc) to various protein substrates in a process called GlcNAcylation. Even though GlcNAc is a common sugar, OGT is the only enzyme that can catalyze its attachment. OGT works by capturing UDP-GlcNAc, a carrier uridine diphosphate (UDP) molecule with GlcNAc, identifying an appropriate target protein, and transferring the GlcNAc from the UDP to a serine or threonine residue on the target protein. A mystery with this system lies with how OGT recognizes the target protein. For the target protein HCF-1, a glutamate residue is located in the position where OGT would normally attach GlcNAc, and OGT will cleave the backbone of HCF-1 instead of transferring a GlcNAc to the target. When the HCF-1glutamate residue is mutated to a serine, OGT will transfer GlcNAc instead of cleaving the backbone. Our project sought to better comprehend the regions that may play an important role in OGT protein recognition using a structure of a portion of HCF-1 fused to OGT. Our model shows that the same five asparagine residues in the tetratricopeptide repeat (TPR) domain of OGT that are important in recognizing traditional target proteins are also involved in recognizing and binding HCF-1, allowing the glutamate residue to bind to the same active site in OGT used for GlcNAcylation.
Wabash College (Crawfordsville IN)
Long time listeners, first time posters ;) Hello from the Wabash College CREST team in Crawfordsville, IN. Team members are Dr. Wally Novak, Roarke Tollar, Titus Edwards, and Nhan Nguyen.
GlmU bifunctional proteins of N-acetyl-D-glucosamine-1-phosphate and uridine-diphosphate-N-acetylglucosamine (GlmU)
PDB file: 2io7 with modifications
Primary Citation: Olsen, Laurence R., et al. “Structure of the E. coli bifunctional GlmU acetyltransferase active site with substrates and products.” Protein Science, vol. 16, no. 6, 2007, pp. 1230–1235., doi:10.1110/ps.072779707.
Abstract: The focus of this project was to explore the structure of the GlmU protein from E. coli and to design and build a physical model that illustrates the key functional features of the protein.
GlmU is a bifunctional bacterial protein that synthesizes uridine-diphosphate-N-acetylglucosamine (UDP-GlcNAc) from D-glucosamine 1-phosphate, acetyl-CoA, and UTP. The structure described here is in complex with the products desulfo-coenzyme A, N-acetyl-D-glucosamine-1-phosphate and UDP-GlcNAc. The N-terminal uridyltransferase domain is composed of two beta-hairpins and a seven-stranded beta-sheet surrounded by alpha-helices. The C-terminal acetyltransferase domain is a left-handed parallel beta-helix. GlmU functions biologically as a homotrimer. The three N-terminal uridyltransferase active sites are independently formed by each monomer; however, each of the three acetyltransferase active sites is comprised of residues from the beta-helix domain of all three monomers. This site is formed by the face and loop region of the first monomer, another face of the second monomer, and the C-terminal tail of the third monomer. This work was funded in part by NSF-DUE 1725940 for the CREST Project.
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Questions about the CREST Program? Contact Margaret Franzen at email@example.com or 414-277-2806. We look forward to hearing from you!