Pan-HSP90 ligand binding reveals isoform-specific differences in plasticity and water networks.

Isoforms of heat shock protein 90 (HSP90) fold oncoproteins that facilitate all 10 hallmarks of cancer. However, its promise as a therapeutic target remains unfulfilled as there is still no FDA-approved drug targeting HSP90 in disease. Among the reasons hindering progress are side effects caused by pan-HSP90 inhibition. Selective targeting of the four isoforms is challenging due to high sequence and structural similarity. Surprisingly, while decades of drug discovery efforts have produced almost 400 human HSP90 structures, no single ligand has been structurally characterized across all four human isoforms to date, which could reveal structural differences to achieve selectivity. To better understand the HSP90 landscape relevant for ligand binding and design we take a three-pronged approach. First, we solved the first complete set of structures of a single ligand bound to all four human isoforms. This enabled a systematic comparison of how side-chains and water networks respond to ligand binding across isoforms. Second, we expanded our analysis to publicly available, incomplete isoform-ligand series with distinct ligand chemistry. This highlighted general trends of protein and water mobility that differ among isoforms and impact ligand binding. Third, we further probed the Hsp90α conformational landscape for accommodating a congeneric series containing the purine scaffold common to HSP90 inhibitors. This revealed how minor ligand modifications flip ligand poses and perturb water and protein conformations. Taken together, this work illustrates how a systematic approach can shed new light on an "old" target and reveal hidden isoform-specific accommodations of congeneric ligands that may be exploited in ligand discovery and design.

Lysyl oxidase in cancer inhibition and metastasis.

Lysyl oxidase is an extracellular matrix, copper - dependent amine oxidase that catalyzes a key enzymatic step in the crosslinking of collagen and elastin. The enzyme is synthesized as a propeptide that is cleaved by procollagen - C - proteinase into two distinct parts: the mature form and the LOX propeptide. The mature enzyme plays a key role in modifying the extracellular matrix and as a result has been implicated in playing a role in the formation of cancer "niches" where tumors will develop and eventually metastasize. On the other hand, the LOX propeptide has been shown to have an inhibitory effect in the development of cancer tumors. New approaches are being developed to test the use of small molecule inhibitors on LOX; however, the lack of a crystal structure has hampered these efforts as it is extremely difficult to design selective inhibitors without knowing what the target receptor looks like. In this mini review we discuss the lysyl oxidase enzyme and its role several types of cancers.

Characterization of an NADH-dependent persulfide reductase from Shewanella loihica PV-4: implications for the mechanism of sulfur respiration via FAD-dependent enzymes.

The NADH-dependent persulfide reductase (Npsr), a recently discovered member of the PNDOR family of flavoproteins that contains both the canonical flavoprotein reductase domain and a rhodanese domain, is proposed to be involved in the dissimilatory reduction of S(0) for Shewanella loihica PV-4. We have previously shown that polysulfide is a substrate for this enzyme, and a recently determined structure of a closely related enzyme (CoADR-Rhod from Bacillus anthracis) suggested the importance of a bound coenzyme A in the mechanism. The work described here shows that the in vivo oxidizing substrates of Npsr are the persulfides of small thiols such as CoA and glutathione. C43S, C531S, and C43,531S mutants were created to determine the role of the flavoprotein domain cysteine (C43) and the rhodanese domain cysteine (C531) in the mechanism. The absolute requirement for C43 in persulfide or DTNB reductase activity shows that this residue is involved in S-S bond breakage. C531 contributes to, but is not required for, catalysis of DTNB reduction, while it is absolutely required for reduction of any persulfide substrates. Titrations of the enzyme with NADH, dithionite, titanium(III), or TCEP demonstrate the presence of a mixed-disulfide between C43 and a tightly bound CoA, and structures of the C43 and C43,531S mutants confirm that this coenzyme A remains tightly bound to the enzyme in the absence of a C43-CoA S-S bond. The structure of Npsr suggests a likely site for binding and reaction with the persulfide substrate on the rhodanese domain. On the basis of kinetic, titration, and structural data, a mechanism for the reduction of persulfides by Npsr is proposed.