ARTICLE | doi:10.20944/preprints202103.0129.v1
Subject: Physical Sciences, Acoustics Keywords: secondary structure elements; nematic order parameter; allostery
Online: 3 March 2021 (11:41:22 CET)
The beautiful structures of single and multi-domain proteins are clearly ordered in some fashion but cannot be readily classified using group theory methods that are successfully used to describe periodic crystals. For this reason, protein structures are considered to be aperiodic, and may have evolved this way for functional purposes, especially in instances that require a combination of softness and rigidity within the same molecule. By analyzing the solved protein structures, we show that orientational symmetry is broken in the aperiodic arrangement of the secondary structural elements (SSEs), which we deduce by calculating the nematic order parameter, $P_2$. We find that the folded structures are nematic droplets with a broad distribution of $P_2$. We argue that non-zero values of $P_2$, leads to an arrangement of the SSEs that can resist mechanical forces, which is a requirement for allosteric proteins. Such proteins, which resist mechanical forces in some regions while being flexible in others, transmit signals from one region of the protein to another (action at a distance) in response to binding of ligands (oxygen, ATP or other small molecules).
REVIEW | doi:10.20944/preprints202002.0099.v1
Subject: Life Sciences, Virology Keywords: retroviruses; HIV-1; reverse transcriptase; mutation rate; drug resistance, allostery
Online: 7 February 2020 (11:50:11 CET)
The high mutation rate of human immunodeficiency virus type 1 (HIV-1) plays a major role in treatment resistance from the development of vaccines to long-lasting drugs. In addressing the crux of the issue, various attempts to estimate the mutation rate of HIV-1 resulted in a large range of 10-5 - 10-3 errors/bp/cycle due to the use of different types of investigation methods. In this review, we discuss the different assay methods, their findings on the mutation rates of HIV-1 and how the location of these mutations can be further analyzed for their potential allosteric effects to reveal potentially new inhibitors with different pharmacodynamics that can be used to circumvent fast occurring HIV drug resistance. Given that HIV is one of the fastest mutating viruses, it is a good model for comprehensive study of its mutations that can give rise to much horizontal understanding towards overall viral drug resistance as well as emerging viral diseases.
ARTICLE | doi:10.20944/preprints201910.0281.v1
Subject: Life Sciences, Biophysics Keywords: protein structural dynamics; NQO1; ligand binding; protein stability; allostery; protein degradation
Online: 24 October 2019 (15:41:46 CEST)
Human NAD(P)H:quinone oxidoreductase 1 (NQO1) is a multi-functional protein whose alteration is associated with cancer, Parkinson´s and Alzheimer´s diseases. NQO1 displays a remarkable functional chemistry, capable of binding different functional ligands that modulate its activity, stability and interaction with proteins and nucleic acids. Our understanding on this functional chemistry is limited by the difficulty of obtaining structural and dynamic information on many of these states. Herein, we have used hydrogen/deuterium exchange monitored by mass-spectrometry (HDXMS) to investigate the structural dynamics of NQO1 in three ligation states: without ligands (NQO1apo), with FAD (NQO1holo) and with FAD and the inhibitor dicoumarol (NQO1dic). We show that NQO1apo has a minimally stable folded core holding the protein dimer and with FAD and dicoumarol ligand binding sites populating binding non-competent conformations. Binding of FAD significantly decreases protein dynamics and stabilizes the FAD and dicoumarol binding sites as well as the monomer:monomer interface. Dicoumarol binding further stabilizes all three functional sites, a result not previously anticipated by available crystallographic models. Our work provides an experimental perspective into the communication of stability effects through the NQO1 dimer, valuable to understand at the molecular level the effects of disease-associated variants, post-translation modifications and ligand binding cooperativity in NQO1.
ARTICLE | doi:10.20944/preprints202110.0178.v1
Subject: Life Sciences, Biochemistry Keywords: cardiac muscle; cardiac contractility; force generation; enhanced molecular dynamics simulations; allostery; protein folding; disease; hypertrophic cardiomyopathy
Online: 12 October 2021 (12:20:26 CEST)
Cardiac myosin-binding protein C (MyBPC) is a thick-filament associated regulatory protein in the sarcomere. It regulates the sensitive contractile system of the myocardium by acting as a mechanical tether, sensitizing the thin filament or modulating myosin motor activity. Mutations in the MYBPC3 gene are a frequent cause for the development of hypertrophic cardiomyopathy, the most frequent cardiac disorder. Recently, the monoallelic double mutation MYBPC3Δ25bp/D389V has been discovered as a subset of the common MYBPC3Δ25bp variant in South Asia. MYBPC3Δ25bp/D389V carriers exhibit hyperdynamic features, which are considered an early finding for the development of hypertrophic cardiomyopathy. Using correlation-guided molecular dynamics simulations sampling, we show that the D389V mutation shifts the conformational distribution of the C2 domain of MyBPC. We further applied biochemical approaches to probe the effects of the D389V mutation on structure, thermostability and protein-protein interactions of MyBPC C2. The melting temperature (Tm) of MyBPC C2 D389V is decreased by 4 to 7 °C compared to wild type while the interaction of the C0-C2 domains with myosin and actin remains unchanged. Additionally, we utilized steered molecular dynamics (SMD) simulations to investigate the altered unfolding pathway of MyBPC C2 D389V. Based on our data, we propose a pathomechanism for the development of HCM in MYBPC3Δ25bp and MYBPC3Δ25bp/D389V carriers.
REVIEW | doi:10.20944/preprints202010.0179.v1
Subject: Life Sciences, Biochemistry Keywords: Non Michaelis-Menten Kinetics; High-throughput screening; allostery; cooperativity; processive kinetics; distributive kinetics; single-molecule; auto-catalytic; drug discovery
Online: 8 October 2020 (13:34:16 CEST)
Biological systems are highly regulated. They are also highly resistant to sudden perturbations enabling them to maintain the dynamic equilibrium essential for sustenance of life. This robustness is conferred by regulatory mechanisms that influence the activity of enzymes/proteins within their cellular context, to adapt to changing environmental conditions. However, the initial rules governing the study of enzyme kinetics were tested and implemented for mostly cytosolic enzyme systems that were easy to isolate and/or recombinantly express. Moreover, these enzymes lacked complex regulatory modalities. Now, with academic labs and pharmaceutical companies turning their attention to more complex systems (for instance, multi-protein complexes, oligomeric assemblies, membrane proteins and post-translationally modified proteins), the initial axioms defined by Michaelis-Menten (MM) kinetics are rendered inadequate and the development of a new kind of kinetic analysis to study these systems is required. The current review strives to present an overview of enzyme kinetic mechanisms that are atypical and, oftentimes, do not conform to the classical MM kinetics. Further, it presents initial ideas on the design and analysis of experiments in early drug-discovery for such systems, to enable effective screening and characterisation of small-molecule inhibitors with desirable physiological outcomes.