Phase Transitions in Small Isotropic Bicelles Erik F. Kot, †,‡ Sergey A. Goncharuk, †,§ Alexander S. Arseniev, †,‡ and Konstantin S. Mineev* ,†,‡ † Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences RAS, str. Miklukho-Maklaya 16/10, Moscow 117997, Russian Federation ‡ Moscow Institute of Physics and Technology, Institutsky per., 9, 141700 Dolgoprudnyi, Russian Federation § Lomonosov Moscow State University, Leninskiye Gory, 1, Moscow 119991, Russian Federation * S Supporting Information ABSTRACT: Isotropic phospholipid bicelles are one of the most prospective membrane mimetics for the structural studies of membrane proteins in solution. Recent works provided an almost full set of data regarding the properties of isotropic bicelles; however, one major aspect of their behavior is still under consideration: the possible mixing between the lipid and detergent in the bilayer area. This problem may be resolved by studying the lipid phase transitions in bicelle particles. In the present work, we investigate two effects: phase transitions of bilayer lipids and temperature-induced growth of isotropic bicelles using the NMR spectroscopy. We propose an approach to study the phase transitions in isotropic bicelles based on the properties of 31 P NMR spectra of bilayer-forming lipids. We show that phase transitions in small bicelles are “fractional”, particles with the liquid-crystalline and gel bilayers coexist in solution at certain temperatures. We study the effects of lipid fatty chain type and demonstrate that the behavior of various lipids in bilayers is reproduced in the isotropic bicelles. We show that the temperature-induced growth of isotropic bicelles is not related directly to the phase transition but is the result of the reversible fusion of bicelle particles. In accordance with our data, rim detergents also have an impact on phase transitions: detergents that resist the temperature-induced growth provide the narrowest and most expressed transitions at higher temperatures. We demonstrate clearly that phase transitions take place even in the smallest bicelles that are applicable for structural studies of membrane proteins by solution NMR spectroscopy. This last finding, together with other data draws a thick line under the long-lasting argument about the relevance of small isotropic bicelles. We show with certainty that the small bicelles can reproduce the most fundamental property of lipid membranes: the ability to undergo phase transition. ■ INTRODUCTION Bicelles are membrane mimetics that are formed in the mixtures of lipids and specific detergents, such as the bile salt derivatives 1 and short-chain phospholipids. 2-4 Bicelles are known to form the discoidal particles, 5 with the lipids forming a patch of planar bilayer and detergents associating into the “rim” of the disc. The size of the particles is controlled by the ratio between the lipid and rim-forming detergent (q) in quite a wide range, starting from 20 kDa. Large bicelles (q >2-3 for various mixtures) were shown to orient spontaneously in the strong magnetic field and are used in solid-state NMR spectroscopy to prepare the oriented bilayer samples 6 and in solution to create the anisotropic environment for the soluble proteins and measure the residual dipolar couplings. 7 Due to the specified ability, such large particles are referred to as “anisotropic” bicelles. In contrast, solutions with small particles which are not capable of spontaneous orientation are called isotropic bicelles (IsoBs). IsoBs have several features that make them an almost perfect membrane mimetic for structural studies of membrane proteins. 8-10 Unlike detergent micelles, bicelles contain lipids that form the plane patch of membrane and properly mimic some properties of the lipid bilayer. 11,12 The lipid composition of bicelles can be varied to simulate the membranes of different cells or their microdomains. 13 Cholesterol, sphingolipids, glycerolipids, lipids with unsaturated fatty chains, or anionic headgroups can be added to the isotropic bicelles. 14-18 IsoBs can be formed using the very mild rim-forming surfactants, such as Facades, 19 that do not cause the unfolding of soluble globular domains of large membrane proteins. 20 Unlike lipid-protein nanodiscs, IsoBs are cheaper and easier to prepare and the size of bicelles is lower, which makes them compatible with solution NMR spectroscopy. They allow the exchange of matter between the particles in solution, which is convenient for the studies of protein-protein interactions. IsoBs have found their application in X-ray crystallography; 21 however, the widest use Received: October 17, 2017 Revised: February 12, 2018 Published: February 27, 2018 Article pubs.acs.org/Langmuir Cite This: Langmuir 2018, 34, 3426-3437 © 2018 American Chemical Society 3426 DOI: 10.1021/acs.langmuir.7b03610 Langmuir 2018, 34, 3426-3437