mPPases create a conserved anionic membrane fingerprint as identified via multi-scale simulations

Abstract

Membrane-integral pyrophosphatases (mPPases) are membrane-bound enzymes responsible for hydrolysing inorganic pyrophosphate and translocating a cation across the membrane. Their function is essential for the infectivity of clinically relevant protozoan parasites and plant maturation. Recent developments have indicated that their mechanism is more complicated than previously thought and that the membrane environment may be important for their function. In this work, we use multiscale molecular dynamics simulations to demonstrate for the first time that mPPases form specific anionic lipid interactions at 4 sites at the distal and interfacial regions of the protein. These interactions are conserved in simulations of the mPPases from Thermotoga maritima, Vigna radiata and Clostridium leptum and characterised by interactions with positive residues on helices 1, 2, 3 and 4 for the distal site, or 9, 10, 13 and 14 for the interfacial site. Due to the importance of these helices in protein stability and function, these lipid interactions may play a crucial role in the mPPase mechanism and enable future structural and functional studies.

Functional and structural asymmetry suggest a unifying principle for catalysis in membrane-bound pyrophosphatases

“…Our model provides, for the first time, an overall explanation of ion selectivity and catalysis in all M-PPases and makes testable predictions: e.g., global conformational changes should occur in the first 0.1–0.2 s at 20 °C. These need to be tested through functional and structural studies, in particular the use of time-resolved, single-molecule techniques to capture further details of mechanism, as well as molecular dynamics simulations—efforts that are already underway (Holmes et al, 2022 ).…”

The inorganic pyrophosphatases of microorganisms: a structural and functional review

“…Further research is needed to correlate the membrane composition of organisms encoding m-PPases, the activity of these enzymes, and their biological role. Holmes, Goldman & Kalli (2022) demonstrated that the lipid environment contributes to intersubunit communication and the current catalytic asymmetry model. For the three proteins studied (from T. maritima , V. radiata, and Clostridium leptum ), the enzymes preferentially interact with anionic lipids rather than with neutral lipids ( Holmes, Goldman & Kalli, 2022 ), which is important given the nature of the organisms that have been found to encode these enzymes, but the environmental conditions under which these organisms thrive are different.…”

“…Computer multiscale simulations have recently addressed the complexity of these enzymes, and the conserved structure found in m-PPases (the number of transmembrane helices and the number of active site structural transitions) indicates that the lipid microenvironment is a determinant of both catalysis and protein dimeric structure ( Holmes, Goldman & Kalli, 2022 ). Further research is needed to correlate the membrane composition of organisms encoding m-PPases, the activity of these enzymes, and their biological role.…”

“…For the three proteins studied (from T. maritima , V. radiata, and Clostridium leptum ), the enzymes preferentially interact with anionic lipids rather than with neutral lipids ( Holmes, Goldman & Kalli, 2022 ), which is important given the nature of the organisms that have been found to encode these enzymes, but the environmental conditions under which these organisms thrive are different. The work by Holmes, Goldman & Kalli (2022) proposes that experimental work is needed to demonstrate the specific interactions with lipids and the residues (that they report are not conserved) present in the protein.…”

Functional and structural asymmetry suggest a unifying principle for catalysis in integral membrane-bound pyrophosphatases

“…global conformational changes should occur in the first 0.1-0.2 s at 20 °C. These need to be tested through functional and structural studies, in particular the use of time-resolved, single-molecule techniques to capture further details of mechanism, as well as efforts to capture mechanistic details through molecular dynamicsefforts that are already underway (Holmes et al, 2022).…”