Exploring the effect of collective mutations in SARS-CoV-2 variants and resistance to antiviral drugs
May 10, 2023
The large number of coronavirus variants is driven by high mutation rate of its RNA genome. As a common target for antiviral drugs, mutations in the Mpro protease are of particular interest. Researchers used the CMCF-BM and SSRL 12-2 beamlines to study the structure of Mpro with a series of naturally occurring mutations for their effect on binding (infectivity) and inhibition (drug resistance). Their results help to understand the mechanisms of antiviral binding and insights into the critical interactions between a virus and its host.Article: https://doi.org/10.1021/acscentsci.3c00054PDBs: 8DJJ, 8DKZ
Novel anticancer drug design starts with understanding how drug-like agonists bind to their target
Feb. 27, 2023
Maintaining mitochondrial proteins is essential to its function as an energy factory. An intricate protein quality control mechanism helps support the mitochondria to control activity and recycle old or misfolded proteins. Interrupting the protein quality control mechanism is a hallmark of many cancers and attractive target for novel drug design.Using the CMCF-BM beamline, researchers from the University of Toronto generated structures of a novel class of analogs which bind and activate the protease ClpP.. Analogs with different affinity to ClpP provide a basis for further modification for rational design of novel anticancer drugs.PDB: 7UVM, 7UVN, 7UVR, 7UVU, 7UW0Reference: Mabanglo, Mark F., et al. "Potent ClpP agonists with anticancer properties bind with improved structural complementarity and alter the mitochondrial N-terminome." Structure (2022). DOI: https://doi.org/10.1016/j.str.2022.12.002See also: CLS News
Understanding the proteins which excrete alginate are made will allow for more targeted treatment of bacterial infection.
Jan. 12, 2023
Biofilms allow microbes to grow on almost any surface and are essential step in most chronic bacterial infections. The basis of biofilms are a complex mixture of polysaccharides with a variety of roles including cell adhesion, motility, and host immune response evasion. Researchers at the SickKids Research Institute used the CMCF-BM beamline to solve the structure of the AlgKX protein complex. Key interactions which directly bind to the alginate polysaccharide were identified and a model for the AlgEKX secretory complex proposed. The research provides a foundation for understanding Pseudomonas is established in chronic cystic fibrosis and novel targets for antibacterial drug design.PDB:: 7ULAGheorghita, A.A., Li, Y.E., Kitova, E.N. et al. Structure of the AlgKX modification and secretion complex required for alginate production and biofilm attachment in Pseudomonas aeruginosa. Nat Commun 13, 7631 (2022). https://doi.org/10.1038/s41467-022-35131-6See also: CLS News
Exploring how coronavirus adapts to new situations...
July 27, 2018
Coronavirus spike protein mediates receptor binding and fusion of viral and host cell membranes. Results from this project provide a model for coronavirus adaptation to environmental changes based on the use of three extended loops for receptor binding, with such loop regions being inherently accommodating to mutational changes. Given their wide host range and zoonotic transmission to humans, coronavirus provides an important model for studying RNA viruses and emergence of new viral threats.PDB ID 6ATK.
How cells overcome some of the toxic effects of benzopyrene...
July 27, 2018
Benzo[a]pyrene (BP) is a frequently-encountered carcinogen that generates a bulky DNA adduct. DNA polymerase kappa is the only known polymerase that can bypass the adduct accurately and protect cells from genotoxic effects. In this Mail-In project, the authors compare binding of normal DNA with BP-adducted DNA to unravel the unique mechanism for accurate replication. PDB ID 4U7C.Nucleic Acids Res. 44, 4957
How bacteria can inactivate certain classes of antibiotics...
July 27, 2018
Macrolides are a class of antibiotic used to treat respiratory tract, skin, and soft tissue infections especially in patients sensitive to β-lactam antibiotics. But, bacteria can inactivate them with macrolide phosphotransferase enzymes. In this structure, azithromycin is bound near the active site of macrolide 2’-phosphotransferase type I, with guanosine. The large antibiotic binding pocket accommodates a variety of macrolides, explaining the broad-spectrum resistance conferred by these enzymes. PDB ID 5IGI.Structure 25, 750
Novel therapeutic targets for cardiac disease and cancer...
July 27, 2018
Protein O-fucosyltransferase 1 (POFUT1) fucosylates cell-surface and secreted glycoproteins including Notch, which is critical during cellular development. Aberrations in this pathway may be linked to generalized Dowling-Degos disease. POFUT1 may also be a novel therapeutic target for cardiac disease and cancer. In this structure obtained from Mail-In data, mouse POFUT1 is in complex with mouse Notch1 EGF26 and GDP. PDB ID 5KY4.Nat. Chem. Biol. 13, 757
How the immune system responds to bacterial infections...
July 27, 2018
LPS is a major bacterial component recognized by the immune system, eliciting an inflammatory response followed by a state of immune tolerance. Acyloxyacyl hydrolase detoxifies LPS to re-establish sensitivity for subsequent infections. The structures described in this work address the questions of LPS recognition and selectivity toward acyl chains. Here, human acyloxyacyl hydrolase is shown with myristic, lauric and palmitic acids hidden deep within a hydrophobic pocket. PDB ID 5W78.Proc. Natl. Acad. Sci. USA 115, E896
Celebrating two milestones reached by scientists who conducted their research at the CMCF
Nov. 23, 2017
CMCF Users have now published 1,000 structures using data collected at the CMCF. There are also 500 peer-reviewed publications acknowledging data collected at the CMCF beamlines. We are pleased to announce that the 1,000th structure is the crystal structure of circumsporozoite protein aTSR domain in complex with 1710 antibody (PDB ID 6B0S, depicted in the image). This structure is part of Jean-Philippe Julien's research into developing a vaccine that prevents the malaria parasite from causing infections. Dr. Julien holds a Canada Research Chair in Structural Immunology and is a scientist in Molecular Medicine at The Hospital for Sick Children Research Institute, as well as assistant professor in the departments of biochemistry and immunology at the University of Toronto. The 500th paper was the result of research by Mirek Cygler's laboratory at the University of Saskatchewan. Using crystallography as well as other techniques, they now have a better understanding of how iron-sulfur clusters are synthesized in the body. These clusters are key components of many proteins critical to life, and defects in the formation of the clusters can cause severe neurological and metabolic diseases, often with fatal outcomes. The findings were published in Nature Communications [Boniecki, MT; Freibert, SA; Mühlenhoff, U; Lill, R; Cygler, M (2017), Structure and functional dynamics of the mitochondrial Fe/S cluster synthesis complex, Nature Communications 8(1)] . Dr. Cygler holds a Canada Research Chair in Molecular Medicine Using Synchrotron Light and is a professor in the department of biochemistry at the University of Saskatchewan.Full CLS News Story
Researchers combine CMCF-BM, CMCF-ID, SAXS & electron microscopy data to discover B-cell CD22 interaction details critical to a healthy immune response...
Oct. 11, 2017
CD22 is a B-cell surface protein involved in regulating the immune response. CD22 knockout mice, for example, have a higher rate of autoimmune disease. CD22's role has been known for some time, but a detailed molecular understanding has been lacking. Now, researchers from the University of Toronto and Hospital for Sick Children (SickKids) have presented a detailed structural model of this key immune component. Combining X-ray crystallography, SAXS and electron microscopy, the researchers describe the molecular structure of human CD22 alone as well as in complex with a sialyllactose ligand or a therapeutic antibody. Initial crystallographic phasing of a portion of CD22 was accomplished by performing a Hg-MAD experiment on beamline CMCF-BM. The resulting structure allowed solution of the structures of the complexes, for which date were obtained on beamline CMCF-ID. SAXS and electron microscopy data rounded out the picture with the result being a description of the full-length extracellular portion of CD22. The work provides key information about the mechanisms controlling B-cell inhibition and clues for designing new autoimmune therapies. PDB ID 5VKJNature Commun. 8, 764
To understand how bacteria can use pili to attach to surfaces, move themselves and become more virulent...
Sept. 13, 2017
Some bacteria are capable of using a "grappling hook" to move themselves. They extend pili to attach to surfaces and retract them to pull themselves toward the point of attachment. In some cases, these pili are essential for the disease process of virulent bacteria. Using data collected at the CMCF, researchers have characterized the "motor" of one such protein system, known as the Type IVa Pilus (T4aP). X-ray crystal structures allowed researchers to deduce a mechanism whereby ATP binds the core ATPase domains, driving the actions of the pili. PDB ID 5TSG.Nature Commun. 8, 15091
Exploring macromolecular machines that may one day produce novel therapeutics...
Jan. 22, 2016
Nonribosomal peptide synthetases (NRPSs) are large, complex proteins responsible for the production of many common antibiotics critical for human health. Making use of the Canadian Macromolecular Crystallography Facility beamlines, researchers have gained a new depth of understanding of NRPSs. Crystal structures of several functional confirmations were determined, supplying critical information about these macromolecular machines. This is an important step if NRPSs are to be used in the production of novel therapeutics. PDB ID: 5es5Nature 529, 239-42
New research describes gut microorganism adaptations to yeast in the diet...
Jan. 20, 2015
The microbial community of the human gut, the microbiota, is critical to human nutrition and health. Different diets are associated with different populations of microbiota. A study has appeared in Nature that explores the adaptation of the microbiota to yeast domestication in the human diet. Yeasts have been an important component of the diet for millenia, through such foods as yeast-leavened breads, fermented beverages and such food products as soy sauce. In this detailed study using multiple techniques, including structural data from the CLS, components of yeast (α-mannans) are shown to be an important food source for Bacteroidetes, a dominant member of the microbiota. These specialized bacteria use a mechanism to break down α-mannans by limited cleavage on the surface, generating large oligosaccharides that are subsequently broken down to mannose by periplasmic enzymes, a process that minimizes nutrient loss. PDBID 4c1r.Nature 517(7533), 165-169
1.0 Å crystal structure of parallel double-stranded poly(A) RNA published...
Aug. 12, 2013
RNA (ribonucleic acid) sequences, although usually single-stranded, can sometimes form double-helical structures. Long RNA sequences having repeating adenine residues, poly(rA), are present on messenger RNA (mRNA), which is transcribed from DNA as a step toward the production of proteins. Poly(rA) RNA was predicted to have the ability to form a double-helix in 1961 based on fibre diffraction experiments. Its detailed structure has only been confirmed by X-ray crystallography recently. Researchers at McGill University have crystallized (rA)11 RNA sequences and combined data collected at the Canadian Light Source and the Cornell High Energy Synchrotron Source to obtain a very detailed 1.0 Å resolution crystal structure of this double-helix. The structure, obtained at physiological pH, shows a parallel double-helix. Ammonium ions stabilize the structure by binding to RNA phosphate groups and adenine N1 atoms, while N7 positions are engaged in hydrogen bonding. Contrary to antiparallel DNA, the poly(rA) double helix shows no major or minor grooves, but rather grooves of equal size. The extent of poly(rA) RNA double-helix formation in mRNA and in other systems remains to be discovered. Researchers believe the structure may be physiologically important, especially under conditions where there is a high local concentration of poly(rA). This can happen, for instance, under conditions where cells are stressed and mRNA become concentrated in RNA granules within cells. PDB ID: 4jrd.Angew. Chem. Int. Ed. Engl., 52, 10370-10373
Antiviral proteins bind foreign RNA within a pocket to detect invaders...
Jan. 23, 2013
During viral infections with viruses such as the flu virus, RNAs having 5'-triphosphate groups (PPP-RNAs) are produced which do not have the usual eukaryotic 5'-cap. A recently-discovered protein, IFIT, can bind this foreign RNA, allowing the immune system to distinguish "self" from "non-self" RNA and initiate processes that serve to prevent the virus from making viable copies of itself. The mechanism for IFIT recognition of foreign viral RNA is the subject of new research, which may pave the way for new developments in the treatment of viral infections. Researchers have used data collected at the CMCF to solve crystal structures of a human IFIT with and without bound PPP-RNAs. These fascinating structures reveal a cavity within the protein designed to accept only single-stranded PPP-RNAs, yet with the necessary ability to be non sequence specific. PDB ID: 4hoqNature 494(7435), 60-64
A struggle to overcome antibiotic resistance using informed design of novel inhibitors...
Sept. 19, 2012
β-lactams are an important class of antibiotics that includes penicillins and carbapenems. A readily-transferable antibiotic resistance factor called New Delhi metallo-β-lactamase-1 (NDM-1) has been found in enteric bacteria. It confers resistance to β-lactams including some critical antibiotics that are presently considered to be the "last line of defence" against multi-drug resistant Gram negative bacteria. The most clinically significant of these lactamases have 2 active site Zn ions. Researchers have used data collected at the CMCF to describe the details of how β-lactam antibiotics are recognized by these Zn-containing enzymes, including a crystal structure having a potential inhibitor bound.J. Am. Chem. Soc. 134(28), 11362-11365
Beamline 08B1-1 plays role in elucidating critical process involved in gene silencing.
Aug. 8, 2012
Small RNA molecules that occur naturally in animals and plants are critical for the regulation of eukaryotic cellular processes. They serve to silence gene expression in various ways including via chromosomal modifications and post-transcriptional effects. These small RNAs are typically 20-30 nucleotides in length and associate with Argonaute proteins to form the RNA-induced silencing complex. In order to function properly, the Argonaute protein must bind to the correct class of small RNA. The 5'-nucleotide of the small RNA is recognized by the MID domain in human Argonaute proteins and this is critical for the correct sorting and association. Now researchers have determined that a similar structural mechanism also occurs in plant Argonaute proteins and, because of the greater complexity of small RNAs in plants, recognition interactions appear to have a corresponding complexity all their own. PDB ID: 4g0x.EMBO J. 31(17), 3588-3595
Some parasites provide proteins to cells that they can later attach to for infiltrating the cells. Researchers examine this process.
Aug. 11, 2011
Toxoplasma and Plasmodium parasites cause numerous diseases worldwide, including malaria and toxoplasmosis. Interestingly, these parasites attack host cells in a very active manner, providing the receptor for binding to the host cell. Interaction thus occurs through a protein called AMA1 to a rhoptry neck (RON) complex provided by the parasite and injected into the host cell. Now researchers have used data collected at the CMCF to determined the structure of AMA1 with a RON2 peptide to give insight into this interaction. PDB ID: 2Y8T and 2Y8S.Science 333, 463-467
Researchers use data from the CMCF to describe the cradle-shaped structure of an unique chaperone protein called Spy.
Feb. 16, 2011
Chaperone proteins make possible the correct folding of other protein molecules in the cell. Researchers have induced bacteria to overproduce a periplasmic chaperone protein called Spy. These unique cradle-shaped dimers help protein refolding and suppress protein aggregation independently of ATP.Nat. Struct. Mol. Biol. 18(3), 262-269
Researchers at U.B.C. make use of beamline 08ID-1 in their study of ryanodine receptor disease hotspots. Mutations in these hotspot regions can contribute to heart and other muscle conditions.
Nov. 27, 2010
Genetic variation in ryanodine receptor 'hotspots' can play a role in diseases affecting muscles, including congenital heart disease. Researchers from the University of British Columbia have combined crystallographic data obtained at beamline 08ID-1 and the Stanford Synchrotron Radiation Lightsource with electron microscopy data to shed light on amino-terminal ryanodine receptor disease hotspot. To view the complete media release click here. PDB ID: 2XOA.Nature 468, 585-588
Some microbes produce enzymes that break down potentially harmful compounds found in pollutants from fossil fuels, plastics or pesticides. Researchers from the University of Guelph make use of data obtained at the CMCF to understand the active site of one such enzyme.
Nov. 8, 2010
HMG/CHA aldolase from Pseudomonas putida is part of a larger pathway for breaking down harmful components of fossil fuel pollution and coal derivatives (fluorene and its analogues) and substances found in plastics and pesticides (phthalate isomers). The researchers have grown crystals of the enzyme and solved the crystal structure in order to better understand how the active site is organized. This has allowed them to propose a catalytic mechanism based on the structural features, kinetics and information available about related aldolases. PDB ID: 3NOJ.J. Biol. Chem. 285, 36608-36615
Malaria, caused by Plasmodia parasites, has re-emerged as a major problem, especially due to multidrug resistance. Researchers from the University of Toronto used data from the 08ID-1 beamline to investigate a promising strategy to develop novel classes of therapeutics.
Aug. 24, 2010
Malaria, caused by Plasmodia parasites, has re-emerged as a major problem, imposing its fatal effects on human health, especially due to multidrug resistance. In Plasmodia, orotidine 5’-monophosphate decarboxylase (ODCase) is an essential enzyme for the de novo synthesis of uridine 5’-monophosphate. Impairing ODCase in these pathogens is a promising strategy to develop novel classes of therapeutics. Researchers from the group of Dr. E. Pai (University of Toronto) used data from the 08ID-1 beamline to investigate the structure–activity relationships of various novel inhibitors of ODCase.J. Med. Chem. 51 (3), 439-448
The Norwalk virus belongs to a superfamily of viruses that stores their genetic code as RNA. Researchers at the University of Calgary used high-resolution data from the 08ID-1 beamline to collect information crucial for drug development.
Aug. 24, 2010
Outbreaks of Norwalk virus are notorious for causing severe dehydration due to vomiting and diarrhea. The currently untreatable bug belongs to a superfamily of viruses that stores their genetic code as RNA. Researchers at the University of Calgary used high-resolution data from the 08ID-1 beamline to determine the structure the Norwalk virus polymerase in various complexed states. The information is crucial to better understand viral replication and for drug development.J. Biol. Chem. 283 (12), 7705-7712
Bacterial infections once thought to be on the verge of eradication have been making a comeback. Researchers at the University of Saskatchewan used the 08ID-1 beamline to study UGM, an enzyme crucial for making a particular "super-antigen", lipopolysaccharide.
Aug. 24, 2010
The need for novel antibiotics is increasingly pressing in the face of the rising threat of bacteria resistant to existing drugs. One approach for such antibiotics is to target the building blocks of bacterial cell walls. One such component is lipopolysaccharide, formed from sugars that are in turn produced using the enzyme UDP-galactopyranose mutase (UGM). Researchers from the University of Saskatchewan isolated UGM in a form bound to sugars and obtained its crystal structure. This information may be used to design drugs that inhibit the enzyme's activity and thus block the formation of bacterial cell walls.J. Mol. Biol. 394 (5), 864-877
University of British Columbia researchers the 08ID-1 beamline to determine the molecular structure of ferritin, an iron storage protein recently discovered in a group of phytoplankton called pennate diatoms.
Aug. 24, 2010
Diatoms are unicellular phytoplankton that account for much of the primary productivity in the world's oceans. The growth and population size of diatoms is dependent on the availability of iron. Using data from the 08ID-1 beamline, researchers from the University of British Columbia determined that pennate diatoms are able to produce an iron-concentrating protein, ferritin, to store iron and thrive in areas that are usually iron-poor.Nature 457 (7228), 467-470
For over 40 years, tuberculosis has been treated using a cocktail of antibiotics that must be taken for six months to a year. A discovery by researchers from the CLS and the University of British Columbia sheds light both on the source of the TB bug’s resilience and a new way to treat the infection
Aug. 24, 2010
Mycobacterium tuberculosis (Mtb), the bacteria that causes tuberculosis, is a resilient organism that can only be effectively treated by a lengthy course of multiple drugs. Mtb is able to survive by harvesting the cholesterol stored in white blood cells. Researchers from the University of British Columbia used the 08ID-1 beamline to collect data about the strucuture of KshAB, one of the enzymes used to break down cholesterol. This structural information can be used to design drugs to interfere with the enzyme and develop improved drugs for the treatment of tuberculosis.Acta Crystallogr. F 64 (9), 805-808