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

The Center for Macromolecular Interactions (CMI) in the department of Biological Chemistry and Molecular Pharmacology at Harvard Medical School has a mission 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)MicroScale Thermophoresis (MST)Differential Scanning Fluorimetry (DSF)Circular Dichroism (CD), and Light Scattering: including size-exclusion chromatography with multi-angle light scattering (SEC-MALS), Dynamic Light Scattering (DLS), and Mass Photometry (MP)

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

Chang S, Thrall ES, Laureti L, Piatt SC, Pagès V, Loparo JJ. Compartmentalization of the replication fork by single-stranded DNA-binding protein regulates translesion synthesis. Nat Struct Mol Biol 2022;29(9):932-941.Abstract
Processivity clamps tether DNA polymerases to DNA, allowing their access to the primer-template junction. In addition to DNA replication, DNA polymerases also participate in various genome maintenance activities, including translesion synthesis (TLS). However, owing to the error-prone nature of TLS polymerases, their association with clamps must be tightly regulated. Here we show that fork-associated ssDNA-binding protein (SSB) selectively enriches the bacterial TLS polymerase Pol IV at stalled replication forks. This enrichment enables Pol IV to associate with the processivity clamp and is required for TLS on both the leading and lagging strands. In contrast, clamp-interacting proteins (CLIPs) lacking SSB binding are spatially segregated from the replication fork, minimally interfering with Pol IV-mediated TLS. We propose that stalling-dependent structural changes within clusters of fork-associated SSB establish hierarchical access to the processivity clamp. This mechanism prioritizes a subset of CLIPs with SSB-binding activity and facilitates their exchange at the replication fork.
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.

CMI News

Refeyn TwoMP instrument for Mass Photometry Installed

November 30, 2022

We are pleased to announce the installation of our newest instrument, the Refeyn TwoMP instrument for Mass Photometry, allowing single-particle mass measurements in solution.  Thanks to the HMS Foundry for helping to fund this new equipment purchase! 

We will begin training in December.  Current CMI users interested in using mass photometry should make a training request in PPMS.  Interested new users should request an account at the CMI.