Periodic Reporting for period 3 - MCS-MD (The Molecular Dynamics of Membrane Contact Sites)
Reporting period: 2022-06-01 to 2023-11-30
Investigation of the mechanisms taking place at MCS has emerged as a central topic in cellular biology in the last few years, and it has led to a large amount of novel cellular, biochemical and structural data that has drastically revolutionized our general understanding of lipid homeostasis in the cell. Yet, due to limitations of experimental methods, a high-resolution understanding of how MCS proteins work is still limited, and the specific molecular details of these mechanisms are still under intense debate, and especially concerning the specificity of lipid transport or the discrimination between lipid sensing and lipid transport.
To understand these processes with unprecedented molecular detail, we have been developing high-throughput protocols based on atomistic and coarse-grain molecular dynamics simulations that leverage and take advantage of all the available, yet often scattered, experimental data. With these approaches, that have not been used so far to investigate MCS because of the extreme complexity of these cellular machineries, we are obtaining a detailed understanding of key molecular processes taking place at MCS, including the specificity of membrane binding, the mechanism of lipid uptake and release, the influence of confinement on protein activity, and the role of membrane lipid composition in the regulation of lipid transport. This approach will drive forward our perception of the limits of structure-based in-silico methods, and it will contribute to our mechanistic understanding of key cellular biology processes by providing new quantitative results that are beyond the current possibilities of experimental approaches.
First, we have extensively investigated and elucidated the mechanism of membrane targeting by MCS proteins. As a first step, we have thoroughly assessed the capability of current all-atom (AA) and coarse-grain (CG) models to investigate peripheral protein-membrane interactions. Next, we have applied the best-identified methods to investigate the association of several MCS protein domains to different model membranes. This detailed molecular understanding of the membrane interface of MCS proteins is a significant progress with respect to current knowledge, as current structural biology approaches often do not provide this piece of information.
Second, we have started investigating the internal dynamics of individual domains of MCS proteins as well as their assemblies. This has helped us characterize the relationship between the dynamics of MCS proteins and their function
Third, we have characterized the mechanism of lipid uptake and release for one specific family of MCS proteins. Our preliminary results recapitulate experimental measurements on transfer rates, and they provide a mechanistic explanation of why lipid transport proteins have a wide range of transport efficiencies.
Fourth, we have worked on improving force field parameters for molecular dynamics simulations in order to correctly reproduce experimental observations. So far, this has led to improved parameters for neutral lipids diacylglycerol and triacylglycerol, but further developments are under way.
In the remaining half of the project, (i) we will continue our development efforts to provide the community with improved technological means to investigate lipid transport at MCS using computer simulations, and (ii) we will establish correlations between MCS protein’s molecular mechanism and their physiological function, in order to elucidate molecular details of cellular lipid homeostasis.