Coronavirus Update

The CMI COVID-19 Plan summarizes the changes implemented at CMI to maintain social distancing, enhance safety and accomodate CMI users. 

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 offers training and access to 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)

The CMI offers data collection services for the characterization of protein secondary structure, mass and oligomeric state, polydispersity, aggregation state, hydrodynamic radius, and thermal stability.  Using a library developed in the lab of Andrew Kruse, the CMI is also offering yeast surface display nanobody selections services.

Recent CMI User Publications

Park EY, Rawson S, Schmoker A, Kim B-W, Oh S, Song KK, Jeon H, Eck M. Cryo-EM structure of a RAS/RAF recruitment complex [Internet]. bioRxiv 2022; Publisher's VersionAbstract
Cryo-EM structures of a KRAS/BRAF/MEK1/14-3-3 complex reveal KRAS bound to the flexible Ras-binding domain of BRAF, captured in two orientations. Autoinhibitory interactions are unperturbed by binding of KRAS and in vitro activation studies confirm that KRAS binding is insufficient to activate BRAF, absent membrane recruitment. These structures illustrate the separability of binding and activation of BRAF by Ras and suggest stabilization of this pre-activation intermediate as an alternative to blocking binding of KRAS.Competing Interest StatementThe authors have declared no competing interest.
Feng J, Dong X, Su Y, Lu C, Springer TA. Monomeric prefusion structure of an extremophile gamete fusogen and stepwise formation of the postfusion trimeric state. Nat Commun 2022;13(1):4064.Abstract
Here, we study the gamete fusogen HAP2 from Cyanidioschyzon merolae (Cyani), an extremophile red algae that grows at acidic pH at 45 °C. HAP2 has a trimeric postfusion structure with similarity to viral class II fusion proteins, but its prefusion structure has been elusive. The crystal structure of a monomeric prefusion state of Cyani HAP2 shows it is highly extended with three domains in the order D2, D1, and D3. Three hydrophobic fusion loops at the tip of D2 are each required for postfusion state formation. We followed by negative stain electron microscopy steps in the process of detergent micelle-stimulated postfusion state formation. In an intermediate state, two or three linear HAP2 monomers associate at the end of D2 bearing its fusion loops. Subsequently, D2 and D1 line the core of a trimer and D3 folds back over the exterior of D1 and D2. D3 is not required for formation of intermediate or postfusion-like states.
Bonazza K, Iacob RE, Hudson NE, Li J, Lu C, Engen JR, Springer TA. Von Willebrand factor A1 domain stability and affinity for GPIbα are differentially regulated by its -glycosylated N- and C-linker. Elife 2022;11Abstract
Hemostasis in the arterial circulation is mediated by binding of the A1 domain of the ultralong protein von Willebrand factor (VWF) to GPIbα on platelets to form a platelet plug. A1 is activated by tensile force on VWF concatemers imparted by hydrodynamic drag force. The A1 core is protected from force-induced unfolding by a long-range disulfide that links cysteines near its N- and C-termini. The O-glycosylated linkers between A1 and its neighboring domains, which transmit tensile force to A1, are reported to regulate A1 activation for binding to GPIb, but the mechanism is controversial and incompletely defined. Here, we study how these linkers, and their polypeptide and O-glycan moieties, regulate A1 affinity by measuring affinity, kinetics, thermodynamics, hydrogen deuterium exchange (HDX), and unfolding by temperature and urea. The N-linker lowers A1 affinity 40-fold with a stronger contribution from its O-glycan than polypeptide moiety. The N-linker also decreases HDX in specific regions of A1 and increases thermal stability and the energy gap between its native state and an intermediate state, which is observed in urea-induced unfolding. The C-linker also decreases affinity of A1 for GPIbα, but in contrast to the N-linker, has no significant effect on HDX or A1 stability. Among different models for A1 activation, our data are consistent with the model that the intermediate state has high affinity for GPIbα, which is induced by tensile force physiologically and regulated allosterically by the N-linker.
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