A sensitive regulator of cellular potassium: A class of potassium channels called K2P channels modulates resting membrane potential in most cells. The channels are regulated by multiple ligands, including the antidepressant drug Prozac, as well as factors such as mechanical stretch and voltage. Dong et al. determined the structure of the human K2P channel, TREK-2, in two conformations and bound to a metabolite of Prozac. The structures show how ligand binding or mechanical stretch might induce switching between the states. Although both states have open channels, one appears primed for gating. A Prozac metabolite binds to the primed state and prevents conformational switching. K2P channels are not a target of Prozac, but their inhibition may contribute to side effects. Science , this issue p. 1256

Lamin Loppers: The nuclear lamina provides mechanical stability to the nuclear envelope and is involved in regulation of cellular processes such as DNA replication. Defects in the nuclear lamina lead to diseases such as progeria and metabolic disorders. One of the components of the nuclear lamina, lamin A, undergoes a complex maturation process. A key player is an inner nuclear membrane zinc metalloprotease (ZMP) that is responsible for two proteolysis steps (see the Perspective by Michaelis and Hrycyna ). Quigley et al. (p. 1604 ) report the crystal structure of human ZMPSTE24 and Pryor et al. (p. 1600 ) that of the yeast homolog Ste24p. The structures provide insight into the mechanism of catalysis and into why mutations in ZMPSTE24 lead to laminopathies.

The regulated movement of glucose across mammalian cell membranes is mediated by facilitative glucose transporters (GLUTs) embedded in lipid bilayers. Despite the known importance of phospholipids in regulating protein structure and activity, the lipid-induced effects on the GLUTs remain poorly understood. We systematically examined the effects of physiologically relevant phospholipids on glucose transport in liposomes containing purified GLUT4 and GLUT3. The anionic phospholipids, phosphatidic acid, phosphatidylserine, phosphatidylglycerol, and phosphatidylinositol, were found to be essential for transporter function by activating it and stabilizing its structure. Conical lipids, phosphatidylethanolamine and diacylglycerol, enhanced transporter activity up to 3-fold in the presence of anionic phospholipids but did not stabilize protein structure. Kinetic analyses revealed that both lipids increase the k_cat of transport without changing the K_m values. These results allowed us to elucidate the activation of GLUT by plasma membrane phospholipids and to extend the field of membrane protein-lipid interactions to the family of structurally and functionally related human solute carriers.

Off-target binding of hydrophobic drugs can lead to unwanted side effects, either through specific or non-specific binding to unintended membrane protein targets. However, distinguishing the binding of drugs to membrane proteins from that of detergents, lipids and cofactors is challenging. Here, we use high-resolution mass spectrometry to study the effects of HIV protease inhibitors on the human zinc metalloprotease ZMPSTE24. This intramembrane protease plays a major role in converting prelamin A to mature lamin A. We monitored the proteolysis of farnesylated prelamin A peptide by ZMPSTE24 and unexpectedly found retention of the C-terminal peptide product with the enzyme. We also resolved binding of zinc, lipids and HIV protease inhibitors and showed that drug binding blocked prelamin A peptide cleavage and conferred stability to ZMPSTE24. Our results not only have relevance for the progeria-like side effects of certain HIV protease inhibitor drugs, but also highlight new approaches for documenting off-target drug binding.

Membrane proteins are essential components of many biochemical processes and are important pharmaceutical targets. Membrane protein structural biology provides the molecular rationale for these biochemical process as well as being a highly useful tool for drug discovery. Unfortunately, membrane protein structural biology is a difficult area of study due to low protein yields and high levels of instability especially when membrane proteins are removed from their native environments. Despite this instability, membrane protein structural biology has made great leaps over the last fifteen years. Today, the landscape is almost unrecognisable. The numbers of available atomic resolution structures have increased 10-fold though advances in crystallography and more recently by cryo-electron microscopy. These advances in structural biology were achieved through the efforts of many researchers around the world as well as initiatives such as the Membrane Protein Laboratory (MPL) at Diamond Light Source. The MPL has helped, provided access to and contributed to advances in protein production, sample preparation and data collection. Together, these advances have enabled higher resolution structures, from less material, at a greater rate, from a more diverse range of membrane protein targets. Despite this success, significant challenges remain. Here, we review the progress made and highlight current and future challenges that will be overcome.

Rationale: Pseudomonas aeruginosa (PA) is an environmental pathogen that commonly infects individuals with cystic fibrosis (CF) and non-CF bronchiectasis, impacting morbidity and mortality. To understand the pathobiology of interactions between the bacterium and host adaptive immunity and to inform rational vaccine design, it is important to understand the adaptive immune correlates of disease. Objectives: To characterize T-cell immunity to the PA antigen outer membrane porin F (OprF) by analyzing immunodominant epitopes in relation to infection status. Methods: Patients with non-CF bronchiectasis were stratified by frequency of PA isolation. T-cell IFN-γ immunity to OprF and its immunodominant epitopes was characterized. Patterns of human leukocyte antigen (HLA) restriction of immunodominant epitopes were defined using HLA class II transgenic mice. Immunity was characterized with respect to cytokine and chemokine secretion, antibody response, and T-cell activation transcripts. Measurements and Main Results: Patients were stratified according to whether PA was never, sometimes (<50%), or frequently (≥50%) isolated from sputum. Patients with frequent PA sputum-positive isolates were more likely to be infected by mucoid PA, and they showed a narrow T-cell epitope response and a relative reduction in Th1 polarizing transcription factors but enhanced immunity with respect to antibody production, innate cytokines, and chemokines. Conclusions: We have defined the immunodominant, HLA-restricted T-cell epitopes of OprF. Our observation that chronic infection is associated with a response of narrowed specificity, despite strong innate and antibody immunity, may help to explain susceptibility in these individuals and pave the way for better vaccine design to achieve protective immunity.