Membrane fluidity adjusts the insertion of the transacylase PlsX to regulate phospholipid biosynthesis in Gram-positive bacteria

Author(s)

Sastre, Diego E. ^[1] ; Basso, Luis G. M. ^[2] ; Trastoy, Beatriz ^[3] ; Cifuente, Javier O. ^[3] ; Contreras, Xabier ^{[4, 5, 6]} ; Gueiros-Filho, Frederico ^[7] ; de Mendoza, Diego ^[8] ; Navarro, Marcos V. A. S. ^[1] ; Guerin, Marcelo E. ^{[4, 3]}

Affiliation

^[1] Univ Sao Paulo, Inst Fis Sao Carlos, Grp Biofis Mol Sergio Mascarenhas, Sao Carlos, SP - Brazil

^[2] Univ Sao Paulo, Fac Filosofia Ciencias & Letras Ribeirao Preto, Dept Fis, Ribeirao Preto, SP - Brazil

^[3] CIC BioGUNE Technol Pk Bizkaia, Struct Biol Unit, Derio 48160, Vizcaya - Spain

^[4] Ikerbasque, Basque Fdn Sci, Bilbao 48013 - Spain

^[5] Univ Basque Country, CSIC, Inst Biofis, Barrio Sarriena S-N, Leioa 48940, Bizkaia - Spain

^[6] Univ Basque Country, Dept Bioquim, Leioa 48940, Bizkaia - Spain

^[7] Univ Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo, SP - Brazil

^[8] Inst Biol Mol & Celular Rosario IBR, Rosario, Santa Fe - Argentina

• Document type: Journal article
• Source: Journal of Biological Chemistry; v. 295, n. 7, p. 2136-2147, FEB 14 2020.
• Web of Science Citations: 2

Abstract

PlsX plays a central role in the coordination of fatty acid and phospholipid biosynthesis in Gram-positive bacteria. PlsX is a peripheral membrane acyltransferase that catalyzes the conversion of acyl-ACP to acyl-phosphate, which is in turn utilized by the polytopic membrane acyltransferase PlsY on the pathway of bacterial phospholipid biosynthesis. We have recently studied the interaction between PlsX and membrane phospholipids in vivo and in vitro, and observed that membrane association is necessary for the efficient transfer of acyl-phosphate to PlsY. However, understanding the molecular basis of such a channeling mechanism remains a major challenge. Here, we disentangle the binding and insertion events of the enzyme to the membrane, and the subsequent catalysis. We show that PlsX membrane binding is a process mostly mediated by phospholipid charge, whereas fatty acid saturation and membrane fluidity remarkably influence the membrane insertion step. Strikingly, the PlsX(L254E) mutant, whose biological functionality was severely compromised in vivo but remains catalytically active in vitro, was able to superficially bind to phospholipid vesicles, nevertheless, it loses the insertion capacity, strongly supporting the importance of membrane insertion in acyl-phosphate delivery. We propose a mechanism in which membrane fluidity governs the insertion of PlsX and thus regulates the biosynthesis of phospholipids in Gram-positive bacteria. This model may be operational in other peripheral membrane proteins with an unprecedented impact in drug discovery/development strategies. (AU)

• FAPESP's process: 15/21583-0 - Development of new antibiotics against super-resistant Staphylococcus aureus strains: identification and characterization of inhibitors of the enzyme Diadenylate cyclase
• Grantee: Marcos Vicente de Albuquerque Salles Navarro
• Support type: Regular Research Grants
• FAPESP's process: 16/05203-5 - HOW DO BACTERIA COORDINATE MEMBRANE BIOGENESIS WITH CELL GROWTH AND DIVISION?
• Grantee: Frederico José Gueiros Filho
• Support type: Regular Research Grants
• FAPESP's process: 14/00206-0 - Structure and function of SARS-CoV spike glycoprotein fusion peptides
• Grantee: Luís Guilherme Mansor Basso
• Support type: Scholarships in Brazil - Post-Doctorate
• FAPESP's process: 14/13411-1 - Study of acyltransferases required for the synthesis of membrane phospholipids in Gram-positive bacteria: an attractive target for antibacterial drug discovery
• Grantee: Diego Emiliano Sastre
• Support type: Scholarships in Brazil - Post-Doctorate