Structural Characterization of Degrader-Induced Ternary Complexes Using Hydrogen-Deuterium Exchange Mass Spectrometry and Computational Modeling: Implications for Structure-Based Design.

The field of targeted protein degradation (TPD) has grown exponentially over the past decade with the goal of developing therapies that mark proteins for destruction leveraging the ubiquitin-proteasome system. One common approach to achieve TPD is to employ a heterobifunctional molecule, termed as a degrader, to recruit the protein target of interest to the E3 ligase machinery. The resultant generation of an intermediary ternary complex (target-degrader-ligase) is pivotal in the degradation process.

A Nanopore Approach for Analysis of Caspase-7 Activity in Cell Lysates.

Caspases are an important protease family that coordinate inflammation and programmed cell death. Two closely related caspases, caspase-3 and caspase-7, exhibit largely overlapping substrate specificities. Assessing their proteolytic activities individually has therefore proven extremely challenging.

Multiple Mechanisms of Zinc-Mediated Inhibition for the Apoptotic Caspases-3, -6, -7, and -8.

Zinc is emerging as a widely used and important biological regulatory signal. Cellular zinc levels are tightly regulated by a complex array of zinc importers and exporters to control processes such as apoptotic cell death. While caspase inhibition by zinc has been reported previously, the reported inhibition constants were too weak to suggest a critical biological role for zinc-mediated inhibition.

Utilizing Inverse Emulsion Polymerization To Generate Responsive Nanogels for Cytosolic Protein Delivery.

Therapeutic biologics have various advantages over synthetic drugs in terms of selectivity, their catalytic nature, and, thus, therapeutic efficacy. These properties offer the potential for more effective treatments that may also overcome the undesirable side effects observed due to off-target toxicities of small molecule drugs. Unfortunately, systemic administration of biologics is challenging due to cellular penetration, renal clearance, and enzymatic degradation difficulties.

Dual Site Phosphorylation of Caspase-7 by PAK2 Blocks Apoptotic Activity by Two Distinct Mechanisms.

Caspases, the cysteine proteases that execute apoptosis, are tightly regulated via phosphorylation by a series of kinases. Although all apoptotic caspases work in concert to promote apoptosis, different kinases regulate individual caspases. Several sites of caspase-7 phosphorylation have been reported, but without knowing the molecular details, it has been impossible to exploit or control these complex interactions, which normally prevent unwanted proliferation.

The facial triad in the α-ketoglutarate dependent oxygenase FIH: A role for sterics in linking substrate binding to O activation.

The factor inhibiting hypoxia inducible factor-1α (FIH) is a nonheme Fe(II)/αKG oxygenase using a 2-His-1-Asp facial triad. FIH activates O via oxidative decarboxylation of α-ketoglutarate (αKG) to generate an enzyme-based oxidant which hydroxylates the Asn residue within the C-terminal transactivation domain (CTAD) of HIF-1α. Tight coupling of these two sequential reactions requires a structural linkage between the Fe(II) and the substrate binding site to ensure that O activation occurs after substrate binds.

Reactive Self-Assembly of Polymers and Proteins to Reversibly Silence a Killer Protein.

Conjugation of biologically active proteins to polymeric materials is of great interest in the treatment of cancer and other diseases of protein deficiency. The conjugation of such biomacromolecules is challenging both due to their hydrophilicity and propensity to denature under non-native conditions. We describe a novel reactive self-assembly approach to "wrap" a protein with polymers, simultaneously protecting its delicate folded state and silencing its enzymatic activity.

A multipronged approach for compiling a global map of allosteric regulation in the apoptotic caspases.

One of the most promising and as yet underutilized means of regulating protein function is exploitation of allosteric sites. All caspases catalyze the same overall reaction, but they perform different biological roles and are differentially regulated. It is our hypothesis that many allosteric sites exist on various caspases and that understanding both the distinct and overlapping mechanisms by which each caspase can be allosterically controlled should ultimately enable caspase-specific inhibition.