Trigger factor both holds and folds its client proteins

ATP-independent chaperones like trigger factor are generally assumed to play passive roles in protein folding by acting as holding chaperones. Here we show that trigger factor plays a more active role. Consistent with a role as an aggregation inhibiting chaperone, we find that trigger factor rapidly binds to partially folded glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and prevents it from non-productive self-association by shielding oligomeric interfaces. In the traditional view of holding chaperone action, trigger factor would then be expected to transfer its client to a chaperone foldase system for complete folding. Unexpectedly, we noticed that GAPDH folds into a monomeric but otherwise rather native-like intermediate state while trigger factor-bound. Upon release from trigger factor, the mostly folded monomeric GAPDH rapidly self-associates into its native tetramer and acquires enzymatic activity without needing additional folding factors. The mechanism we propose here for trigger factor bridges the holding and folding activities of chaperone function

A bovine antibody possessing an ultralong complementarity-determining region CDRH3 targets a highly conserved epitope in sarbecovirus spike proteins

Broadly neutralizing antibodies (bNAbs) have huge potential as novel anti-viral therapeutics due to their ability to recognize highly conserved epitopes that are seldom mutated in viral variants. A sub-set of bovine antibodies possess an ultralong complementarity-determining region (CDR)H3 that is highly adept at recognizing such conserved epitopes, but their reactivity against Sarbecovirus Spike proteins has not been explored previously. Here, we use a SARS-naïve library to isolate a broadly reactive bovine CDRH3 that binds the receptor-binding domain of SARS-CoV, SARS-CoV-2, and all SARS-CoV-2 variants. We show further that it neutralizes viruses pseudo-typed with SARS-CoV Spike, but this is not by competition with angiotensin-converting enzyme 2 (ACE2) binding. Instead, using differential hydrogen-deuterium exchange mass spectrometry, we demonstrate that it recognizes the major site of vulnerability of Sarbecoviruses. This glycan-shielded cryptic epitope becomes available only transiently via interdomain movements of the Spike protein such that antibody binding triggers destruction of the prefusion complex. This proof of principle study demonstrates the power of in vitro-expressed bovine antibodies with ultralong CDRH3s for the isolation of novel, broadly reactive tools to combat emerging pathogens and to identify key epitopes for vaccine development