In-solution and interface study on biological model membranes and their interaction with Prohibitin

Piccinini, Alice

Abstract

Prohibitins (PHB) play a crucial role in premature cellular ageing, tumour suppression, cell cycle regulation, and apoptosis, and their deregulation is involved in viral infections, neurological and cardiovascular diseases and cancer. Despite the essential role of this complex, little is known regarding its structure.
PHB are highly conserved heterodimeric proteins composed of two subunits, PHB1 and PHB2, which are known to self-organize as a multimeric (16-20 individual proteins) ring at the inner mitochondrial membrane. Each PHB subunit has an N-terminal region that anchors the prohibitin complex to the membrane.
This thesis aims to characterize the interaction between the N-terminal portion of PHB with the membrane and investigate a possible synergy between the two PHB homologues. Furthermore, this thesis aims to understand the role of cardiolipin in the interaction of PHB with the membrane. This is supported by several studies that report cardiolipin interacting with Prohibitin to maintain the asymmetric lipid distribution of the mitochondrial membrane.
Using interface techniques such as Quartz-Crystal-Microbalance with dissipation monitoring, neutron, and X-ray reflectometry and bulk techniques on liposomes such as small-angle neutron and X-ray scattering, we investigated synthetic prohibitin peptides of the length of 20 (PHB1) and 24 (PHB2) amino acids in model membranes.
Our results revealed that the N-terminal PHB peptides disrupt the membrane microstructure, demonstrating a degree of synergism, but did not show a strong membrane affinity. The presence of cardiolipin and/or phosphatidylethanolamine enhances the effect of the peptides on membranes and induces a modification of the membrane microstructure. We show that the effect of cardiolipin on the membrane is dependent on both cardiolipin content as well as the overall membrane composition.
In addition, we conducted a lipidomic study on the thermophile fungus, C. thermophilum, to investigate the membrane composition in mitochondria versus total lipid extract, which may give clues as to the temperature resistance of thermophilic organisms. Our preliminary results are in line with published literature, however, phosphatidylserine was detected in total lipid extract only.
In this thesis, we examined in detail the role of membrane composition with regard to membrane microstructure, vesicle morphology, peptide insertion and potential temperature resistance. These are important aspects with respect to organellespecific membrane composition and targeted protein interactions in membranes.

Citation

Piccinini, A. (2023). In-solution and interface study on biological model membranes and their interaction with Prohibitin. (Thesis). Keele University

• Thesis Type: Thesis
• Deposit Date: Sep 27, 2023
• Additional Information: Embargo on access until 31 July 2026 - The thesis is due for publication, or the author is actively seeking to publish this material.
• Award Date: 2023-07