Membranes constitute a meeting point for lipids and proteins. Not only do they define the entity of cells and cytosolic organelles but they also display a wide variety of important functions previously ascribed to the activity of proteins alone. Indeed, lipids have commonly been considered a mere support for the transient or permanent association of membrane proteins, while acting as a selective cell/organelle barrier. However, mounting evidence demonstrates that lipids themselves regulate the location and activity of many membrane proteins, as well as defining membrane microdomains that serve as spatio-temporal platforms for interacting signalling proteins. Membrane lipids are crucial in the fission and fusion of lipid bilayers and they also act as sensors to control environmental or physiological conditions. Lipids and lipid structures participate directly as messengers or regulators of signal transduction. Moreover, their alteration has been associated with the development of numerous diseases. Proteins can interact with membranes through lipid co-/post-translational modifications, and electrostatic and hydrophobic interactions, van der Waals forces and hydrogen bonding are all involved in the associations among membrane proteins and lipids. The present study reviews these interactions from the molecular and biomedical point of view, and the effects of their modulation on the physiological activity of cells, the aetiology of human diseases and the design of clinical drugs. In fact, the influence of lipids on protein function is reflected in the possibility to use these molecular species as targets for therapies against cancer, obesity, neurodegenerative disorders, cardiovascular pathologies and other diseases, using a new approach called membrane-lipid therapy.

TRPV1 (transient receptor potential vanilloid receptor subtype 1) is a member of the TRP channel family gated by vanilloids, protons, and heat. Structurally, TRPV1 appears to be a tetramer formed by the assembly of four identical subunits around a central aqueous pore. The molecular determinants that govern its subunit oligomerization remain elusive. Here, we report the identification of a segment comprising ⁶⁸⁴Glu-⁷²¹Arg (referred to as the TRP-like domain) in the C terminus of TRPV1 as an association domain (AD) of the protein. Purified recombinant C terminus of TRPV1 (TRPV1-C) formed discrete and stable multimers in vitro. Yeast two-hybrid and pull-down assays showed that self-association of the TRPV1-C is blocked when segment ⁶⁸⁴Glu-⁷²¹Arg is deleted. Biochemical and immunological analysis indicate that removal of the AD from full-length TRPV1 monomers blocks the formation of stable heteromeric assemblies with wild-type TRPV1 subunits. Deletion of the AD in a poreless TRPV1 subunit suppressed its robust dominant-negative phenotype. Together, these findings are consistent with the tenet that the TRP-like domain in TRPV1 is a molecular determinant of the tetramerization of receptor subunits into functional channels. Our observations suggest that the homologous TRP domain in the TRP protein family may function as a general, evolutionary conserved AD involved in subunit multimerization.

The rapid advances of 3D techniques for the structural determination of proteins and the development of numerous computational methods and strategies have led to identifying highly active compounds in computer drug design. Molecular docking is a method widely used in high-throughput virtual screening campaigns to filter potential ligands targeted to proteins. A great variety of docking programs are currently available, which differ in the algorithms and approaches used to predict the binding mode and the affinity of the ligand. All programs heavily rely on scoring functions to accurately predict ligand binding affinity, and despite differences in performance, none of these docking programs is preferable to the others. To overcome this problem, consensus scoring methods improve the outcome of virtual screening by averaging the rank or score of individual molecules obtained from different docking programs. The successful application of consensus docking in high-throughput virtual screening highlights the need to optimize the predictive power of molecular docking methods.

Transient receptor potential vanilloid receptor subtype I (TRPV1) is an ion channel gated by physical and chemical stimuli that belongs to the TRPV protein family. TRPV receptors contain a highly conserved, 6‐mer segment near the channel gate, known as the TRP box, whose function remains unknown. Here, we performed an alanine scanning mutagenesis of the TRP box of TRPV1 (IWKLQR) and found that mutation of this motif affected channel gating by raising the free energy of channel activation. Functional characterization of TRPV1 mutants showed that substitution of I696, W697, and R701 by alanine severely affected voltage‐ and heat‐dependent activation and notably reduced the capsaicin responsiveness and tachyphylaxia, while mutation of K698, L699, and Q700 had minor effects. In addition, mutation of I696 to alanine promoted a strong outward rectification at negative membrane potentials, and slowed the kinetics of channel activation. Taken together, our findings suggest that modification of I696, W697, and R701 to alanine altered channel function by affecting events downstream of the initial stimuli‐sensing step and imply that inter‐subunit interactions within the TRP box play an important role in TRPV1 gating.—Valente, P., García‐Sanz, N., Gomis, A., Fernández‐Carvajal, A., Fernández‐Ballester, G., Viana, F., Belmonte, C., and Ferrer‐Montiel, A. Identification of molecular determinants of channel gating in the transient receptor potential box of vanilloid receptor I. FASEB J. 22, 3298–3309 (2008)

Transient receptor potential vanilloid receptor subtype 1 (TRPV1) is an ionotropic receptor activated by temperature and chemical stimuli. The C-terminal region that is adjacent to the channel gate, recognized as the TRP domain, is a molecular determinant of receptor assembly. However, the role of this intracellular domain in channel function remains elusive. Here, we show that replacement of the TRP domain of TRPV1 with the cognate region of TRPV channels (TRPV2–TRPV6) did not affect receptor assembly and trafficking to the cell surface, although those receptors containing the TRP domain of the distantly related TRPV5 and TRPV6 did not display ion channel activity. Notably, functional chimeras exhibited an impaired sensitivity to the activating stimuli, consistent with a significant contribution of this protein domain to channel function. At variance with TRPV1, voltage-dependent gating of chimeric channels could not be detected in the absence of capsaicin and/or heat. Biophysical analysis of functional chimeras revealed that the TRP domain appears to act as a molecular determinant of the activation energy of channel gating. Together, these findings uncover a role of the TRP domain in intersubunit interactions near the channel gate that contribute to the coupling of stimulus sensing to channel opening.