Berger, B. W. | bioRxiv Science

bioRxiv2023

Structural analyses of 'substrate-pH of activity' pairing observed in Polysaccharide lyases

Anionic polysaccharides found in nature are functionally and structurally diverse, and so are the polysaccharide lyases (PLs) which catalyse their degradation. Atomic superposition of various PL folds according to their cleavable substrate structure confirm the occurrence of structural convergence at PL active sites. This suggests that various PL folds have emerged to cleave a particular class of anionic polysaccharide during the course of evolution. While structural and mechanistic similarity of PLs active site have been highlighted in earlier studies, a detailed understanding regarding functional properties of this catalytic convergence remains an open question, especially the role of extrinsic factors such as pH in the context of substrate binding and catalysis. Our earlier structural and functional work on pH directed multi-substrate specificity of Smlt1473 inspired us to regroup PLs according to substrate type to analyse the pH dependence of their catalytic activity. Interestingly, we find that particular groups of substrates are cleaved in a particular pH range (acidic/neutral/basic) irrespective of PLs fold, boosting the idea of functional convergence as well. Based on this observation, we set out to define structurally and computationally the key constituents of an active site among PL families. This study delineates the structural determinants of conserved substrate-pH activity pairing within and between PL families.

biochemistry↗

bioRxiv2019

Disrupting the Transmembrane Domain Oligomerization of Protein Tyrosine Phosphatase Receptor J Promotes its Activity in Cancer Cells

Despite the critical regulatory roles that receptor protein tyrosine phosphatases (RPTP) play in mammalian signal transduction, the detailed structural basis for the regulation of their catalytic activity is not fully understood, nor are they generally therapeutically targetable. It is due, in part, to the lack of known natural ligands or selective agonists. In contrast to conventional structure-function relationship for receptor tyrosine kinases (RTKs), the activity of RPTPs has been reported to be suppressed by dimerization, which may prevent their access to their RTK substrates. We report here that: (i) homodimerization of PTPRJ (also known as DEP-1) is regulated by specific transmembrane (TM) residues, and (ii) disrupting these interactions can destabilize full-length PTPRJ homodimerization in cells, reduce the phosphorylation of EGFR (a known substrate) and downstream signaling effectors, antagonize EGFR-driven cell phenotypes, and promote substrate access. We demonstrate these points in human cancer cells using both mutational studies and through the identification of a peptide designed to bind to the PTPRJ TM domain. This peptide is the first example of such allosteric agonist of RPTPs. This study, therefore, provides not only fundamental structure-function insights on how PTPRJ activity is tuned by TM interactions in cells but also opportunities to develop a unique class of agents that could be used as tools to probe RPTPs signaling regulating mechanisms or for therapeutic purposes in cancers driven by RTK signaling.

biochemistry↗