Positive cooperativity in the activation of E. coli aquaporin Z by cardiolipin: potential for lipid-based aquaporin modulators

Abstract

Aquaporins (AQPs) are tetrameric integral membrane proteins (IMPs) that permeabilize cellular membranes to water [1]. Their architecture is well conserved from bacteria to humans [2] where each monomer (~ 25 kDa) acts as a water channel [3]. Although AQPs in humans regulate water transport in several organs and their modulation has immense therapeutic potential, high-throughput screening campaigns to search for AQP inhibitors and modulators have been mostly fruitless [4]. Recently, a new avenue to AQP modulation was identified using native mass spectrometry in the gas phase when E. coli cardiolipin (eCDL) was found to bind E. coli aquaporin Z (AqpZ) [5]. The same paper showed that the activity of AqpZ was reduced to less than half when reconstituted in E. coli lipids (ECL) that did not have eCDL. These findings are not entirely surprising since CDL is a four-fatty acid anionic lipid present in both mammalian and bacterial membranes that is known to bind to several membrane proteins with a stabilizing and/or activating effect [6–9]. However, although it has been predicted that eCDL binds at the monomer-monomer interfaces of the AqpZ tetramer [5,10], neither binding site, stoichiometry or precise mode of activation of AqpZ by eCDL are known. The bulk lipid in E. coli is zwitterionic phosphatidylethanolamine (PE) (60%), followed by anionic phosphatidyl-glycerol (PG) (15%), eCDL (5–15%) and other minor contributors [11]. The last two, eCDL and PG have been suggested to be present in the annular regions around AqpZ, whereas PE forms the bulk of the membrane [10]. Herein, we have studied the role of PG and eCDL in the modulation of AqpZ proteoliposome permeability.