• center for macromolecular interactions
  • CMI Laboratory

    Welcome to the CMI

    An HMS core for the characterization of macromolecules and their interactions.

  • CMI SEC-MALS system

    CMI SEC-MALS system

    Multi-Angle Light Scattering and Dynamic Light Scattering are used to measure the molar mass and hydrodynamic radius.

  • CMI Jasco J815 CD instrument

    CMI Spectropolarimeter

    Circular Dichroism is used to examine protein secondary structure in solution.

  • CMI MST and ITC instruments

    CMI MST and ITC Instruments

    Isothermal Titration Calorimetry and Microscale Thermophoresis measuring binding in solution.

  • CMI SPR and BLI instruments

    CMI SPR and BLI Instruments

    Surface Plasmon Resonance and Biolayer Interferometry are used to measure binding kinetics.

Center for Macromolecular Interactions

Welcome to the Center for Macromolecular Interactions (CMI) in the department of Biological Chemistry and Molecular Pharmacology at Harvard Medical School.  Our mission is to enhance basic research in the HMS community by providing scientific consultation, training and access to shared biophysical instruments for the characterization and analysis of macromolecules and their complexes.

The facility currently includes instruments for Isothermal Titration Calorimetry (ITC)Surface Plasmon Resonance (SPR)Biolayer Interferometry (BLI)Differential Scanning Fluorimetry (DSF)Circular Dichroism (CD)Light Scattering: size-exclusion chromatography with multi-angle light scattering (SEC-MALS) and Dynamic Light Scattering (DLS), and MicroScale Thermophoresis (MST)

Recent User Publications

Bernard-Gauthier V, Mossine AV, Knight A, Patnaik D, Zhao W, Cheng C, Krishnan HS, Xuan LL, Chindavong PS, Reis SA, Chen JM, Shao X, Stauff J, Arteaga J, Sherman PS, Salem N, Bonsall D, Amaral B, Varlow C, Wells L, Martarello L, Patel S, Liang SH, Kurumbail RG, Haggarty SJ, Scott PJH, Vasdev N. Structural Basis for Achieving GSK-3 Inhibition with High Potency, Selectivity and Brain Exposure for PET Imaging and Drug Discovery. J Med Chem 2019;Abstract
Using structure-guided design, several cell based assays, and microdosed positron emission tomography (PET) imaging, we identified a series of highly potent, selective and brain penetrant oxazole-4-carboxamide-based inhibitors of glycogen synthase kinase-3 (GSK-3). An isotopolog of our first-generation lead, [3H]PF-367, demonstrates selective and specific target engagement in vitro, irrespective of activation state. We discovered substantial ubiquitous GSK-3-specific radioligand binding in Tg2576 Alzheimer's disease (AD), suggesting application for these compounds in AD diagnosis, and identified [11C]OCM-44 as our lead GSK-3 radiotracer, with optimized brain uptake by PET imaging in nonhuman primates. GSK-3-isozyme selectivity was assessed to reveal OCM-51, the most potent (IC50 = 0.030 nM) and selective (>10-fold GSK-3/GSK-3) GSK-3 inhibitor known to date. Inhibition of CRMP2T514 and tau phosphorylation, as well as favorable therapeutic window against WNT/-catenin signaling activation was observed in cells.
Rovere M, Powers AE, Jiang H, Pitino JC, Fonseca-Ornelas L, Patel DS, Achille A, Langen R, Varkey J, Bartels T. E46K-like α-synuclein mutants increase lipid interactions and disrupt membrane selectivity. J Biol Chem 2019;294(25):9799-9812.Abstract
Parkinson's disease (PD) is one of the most common neurodegenerative disorders, and both genetic and histopathological evidence have implicated the ubiquitous presynaptic protein α-synuclein (αSyn) in its pathogenesis. Recent work has investigated how disrupting αSyn's interaction with membranes triggers trafficking defects, cellular stress, and apoptosis. Special interest has been devoted to a series of mutants exacerbating the effects of the E46K mutation (associated with autosomal dominant PD) through homologous Glu-to-Lys substitutions in αSyn's N-terminal region ( E35K and E61K). Such E46K-like mutants have been shown to cause dopaminergic neuron loss and severe but L-DOPA-responsive motor defects in mouse overexpression models, presenting enormous translational potential for PD and other "synucleinopathies." In this work, using a variety of biophysical techniques, we characterize the molecular pathology of E46K-like αSyn mutants by studying their structure and membrane-binding and remodeling abilities. We find that, although a slight increase in the mutants' avidity for synaptic vesicle-like membranes can be detected, most of their deleterious effects are connected to their complete disruption of αSyn's curvature selectivity. Indiscriminate binding can shift αSyn's subcellular localization away from its physiological interactants at the synaptic bouton toward trafficking vesicles and organelles, as observed in E46K-like cellular and murine models, as well as in human pathology. In conclusion, our findings suggest that a loss of curvature selectivity, rather than increased membrane affinity, could be the critical dyshomeostasis in synucleinopathies.
Watanabe A, McCarthy KR, Kuraoka M, Schmidt AG, Adachi Y, Onodera T, Tonouchi K, Caradonna TM, Bajic G, Song S, McGee CE, Sempowski GD, Feng F, Urick P, Kepler TB, Takahashi Y, Harrison SC, Kelsoe G. Antibodies to a Conserved Influenza Head Interface Epitope Protect by an IgG Subtype-Dependent Mechanism. Cell 2019;177(5):1124-1135.e16.Abstract
Vaccines to generate durable humoral immunity against antigenically evolving pathogens such as the influenza virus must elicit antibodies that recognize conserved epitopes. Analysis of single memory B cells from immunized human donors has led us to characterize a previously unrecognized epitope of influenza hemagglutinin (HA) that is immunogenic in humans and conserved among influenza subtypes. Structures show that an unrelated antibody from a participant in an experimental infection protocol recognized the epitope as well. IgGs specific for this antigenic determinant do not block viral infection in vitro, but passive administration to mice affords robust IgG subtype-dependent protection against influenza infection. The epitope, occluded in the pre-fusion form of HA, is at the contact surface between HA head domains; reversible molecular "breathing" of the HA trimer can expose the interface to antibody and B cells. Antigens that present this broadly immunogenic HA epitope may be good candidates for inclusion in "universal" flu vaccines.
Rogers JM, Waters CT, Seegar TCM, Jarrett SM, Hallworth AN, Blacklow SC, Bulyk ML. Bispecific Forkhead Transcription Factor FoxN3 Recognizes Two Distinct Motifs with Different DNA Shapes. Mol Cell 2019;74(2):245-253.e6.Abstract
Transcription factors (TFs) control gene expression by binding DNA recognition sites in genomic regulatory regions. Although most forkhead TFs recognize a canonical forkhead (FKH) motif, RYAAAYA, some forkheads recognize a completely different (FHL) motif, GACGC. Bispecific forkhead proteins recognize both motifs, but the molecular basis for bispecific DNA recognition is not understood. We present co-crystal structures of the FoxN3 DNA binding domain bound to the FKH and FHL sites, respectively. FoxN3 adopts a similar conformation to recognize both motifs, making contacts with different DNA bases using the same amino acids. However, the DNA structure is different in the two complexes. These structures reveal how a single TF binds two unrelated DNA sequences and the importance of DNA shape in the mechanism of bispecific recognition.
Pitts J, Hsia C-Y, Lian W, Wang J, Pfeil M-P, Kwiatkowski N, Li Z, Jang J, Gray NS, Yang PL. Identification of small molecule inhibitors targeting the Zika virus envelope protein. Antiviral Res 2019;164:147-153.Abstract
The recent emergence of Zika virus, a mosquito-borne flavivirus, in the Americas has shed light on the severe neurological diseases associated with infection, notably congenital microcephaly in newborns and Guillain-Barré syndrome in adults. Despite the recent focus on Zika virus, there are currently no approved vaccines or antiviral therapies available to treat or prevent infection. In this study we established a competitive amplified luminescent proximity homogeneous assay (ALPHAscreen) to identify small molecule inhibitors targeting the envelope protein of Zika virus (Zika E). We utilized this assay to screen two libraries of nearly 27,000 compounds and identified seven novel inhibitors of Zika E. Characterization of these primary screening leads demonstrated that inhibition of Zika virus occurs at non-cytotoxic concentrations for all seven lead compounds. In addition, we found that all seven lead compounds have potent activity against the closely related dengue virus 2 but not vesicular stomatitis virus, an unrelated enveloped virus. Biochemical experiments indicate that these compounds act by preventing E-mediated membrane fusion. This work highlights a new method for the discovery and optimization of direct-acting antivirals targeting the E protein of Zika and other flaviviruses.
More

CMI News

New Instruments and Services coming coon!

November 5, 2019
Thanks to generous funding from the HMS Foundry, the CMI will be adding new instrumentation and new services in the coming months.  Stay tuned for more information.

Wyatt WISH module

July 30, 2019
We've added a Wyatt WISH module (1 ml high-pressure injection system) to our SEC-MALS capabilities.  This is particularly useful for batch measurements such as and dn/dc measurements. Contact cmi@hms.harvard.edu to learn more or to try it out. 
More