BMOL Seminar Series: Dr. Dominique Frueh

Seminar Details

Speaker: Dr. Dominique Frueh, Associate Professor of Biophysics and Biophysical Chemistry at Johns Hopkins University

Host: Dr. Rajesh Nagarajan, Chemistry & Biochemistry

Title: When binding is not simple: NMR reveals the prominence of structural dynamics in peptide synthetase domain engagement.

Abstract: In this presentation, I will illustrate how the non-invasive nature of NMR can be harnessed to probe molecular responses in macromolecules during biochemical reactions and identify and determine transient states in macromolecules. I will first explain how NMR helps us understand domain communication in nonribosomal peptide synthetases, and later describe NMR methods we developed to overcome challenges in NMR studies of larger proteins. Nonribosomal peptide synthetases (NRPSs) use repeats of domains to tether substrates onto carrier protein domains (CP) and assemble them into complex products through intervening condensation or cyclization domains. A dynamic domain architecture has hampered NRPS engineering because it is unclear whether and how catalytic steps required for synthesis drive transient domain interactions. Here, we demonstrate experimentally that large-scale structural dynamics within a 52 kDa cyclization domain sense the attachment of substrates to its partner CP and promote binding while opening an allosteric path to a remote binding site. The demonstration relies on a nuclear magnetic resonance experimental design, in which we monitor the molecular response of a protein towards covalent modifications of its binding partner in situ. Further, I will discuss how we demonstrated that carrier proteins interact with their prosthetic groups and do not simply serve as an anchor for substrates. Assigning the NMR resonances of a 453-residue protein is challenging, and time permitting, I will show how mathematical operations applied to NMR spectra can provide new correlation maps that facilitate their analysis and how NMR can help optimize complex chemo-enzymatic reactions. Our results exemplify how structural dynamics within proteins can couple substrate recognition with active site remodeling and remote protein communication, and they illustrate the versatility of NMR as an atomic readout of biomolecular function.

Publication Related to Talk: Global protein dynamics as communication sensors in peptide synthetase domains