Biology subjects

Lake, E. W.

Publications and source records attributed to Lake, E. W..

2 recordsLinked to original sources

bioRxiv2024

The structural basis for the selective antagonism of soluble TNF-alpha by shark variable new antigen receptors

The pro-inflammatory cytokine tumor necrosis factor-alpha (TNF)- is synthesized as transmembrane TNF- (tmTNF-) where proteolytic processing releases soluble TNF- (sTNF-). tmTNF- can act as either a ligand by activating TNF receptors, or a receptor that transmits outside-to-inside signals (reverse signalling) after binding to native receptors. All TNF- therapies bind tmTNF- and induce reverse signalling which can result in immunosuppression leading to infection. We present crystal structures of two anti-TNF- Variable New Antigen Receptors (VNAR) in complex with sTNF- via two distinct epitopes. The VNAR-D1 recognized an epitope that selectively engaged sTNF- while VNAR-C4 bound an epitope that overlapped with other biologic therapies. In activated CD4+ T cells, our VNARs did not bind tmTNF- in contrast to commercially available therapies that demonstrated induction of reverse signalling. Our findings suggest that neutralisation through a unique mechanism may lead to anti-TNF- agents with an improved safety profile that will benefit high-risk patients.

pharmacology and toxicology↗

bioRxiv2017

A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation

Many eukaryotic protein kinases are activated by phosphorylation on a specific conserved residue in the regulatory activation loop, a post-translational modification thought to stabilize the active DFG-In state of the catalytic domain. Here we use a battery of spectroscopic methods that track different catalytic elements of the kinase domain to show that the ~100-fold activation of the mitotic kinase Aurora A (AurA) by phosphorylation occurs without a population shift to the DFG-In state, and that the activation loop of the activated kinase remains highly dynamic. Instead, molecular dynamics simulations and electron paramagnetic resonance experiments show that phosphorylation profoundly alters the structure and dynamics of the DFG-In subpopulation, leading to activation of the kinase. Kinetics experiments tracking structural transitions during nucleotide binding suggest that a substantial DFG-Out subpopulation is an important feature of activated AurA that evolved to optimize the kinetics of substrate binding and product release.

biophysics↗