Examples abound of membrane-bound enzymes for which the local membrane environment plays an important role, including the ectoenzyme that triggers blood clotting, the plasma serine protease, factor VIIa, bound to the integral membrane protein, tissue factor. The activity of this enzyme complex is markedly influenced by lipid bilayer composition and further by tissue factor partitioning into membrane microdomains on some cell surfaces. Unfortunately, little is known about how membrane microdomain composition controls factor VIIa-tissue factor activity, as reactions catalyzed by membrane-tethered enzymes are typically studied under conditions in which the experimenter cannot control the composition of the membrane in the immediate vicinity of the enzyme. To overcome this problem, we used a nanoscale approach that afforded complete control over the membrane environment surrounding tissue factor by assembling the factor VIIa⅐tissue factor complex on stable bilayers containing 67 ؎ 1 phospholipid molecules/leaflet (Nanodiscs). We investigated how local changes in phospholipid bilayer composition modulate the activity of the factor VIIa⅐tissue factor complex. We also addressed whether this enzyme requires a pool of membrane-bound protein substrate (factor X) for efficient catalysis, or alternatively if it could efficiently activate factor X, which binds directly to the membrane nanodomain adjacent to tissue factor. We have shown that full proteolytic activity of the factor VIIa⅐tissue factor complex requires extremely high local concentrations of anionic phospholipids and further that a large pool of membrane-bound factor X is not required to support sustained catalysis.In both normal hemostasis and many life-threatening thrombotic diseases, blood clotting is triggered when tissue factor (TF), 5 an integral membrane protein, binds the plasma serine protease factor VIIa (fVIIa) (1). The resulting two-subunit membrane-bound enzyme TF⅐VIIa activates the plasma zymogen factors IX (fIX) and X (fX) by limited proteolysis. Erwin Chargaff (of nucleic acids fame) demonstrated in the 1940s that TF procoagulant activity requires it to be associated with phospholipids, and research over the ensuing decades has demonstrated that TF is a membrane-spanning protein that must be incorporated into bilayers containing anionic phospholipids for optimal activity (reviewed by Bach (2)). Despite this extensive history, we still have an indistinct picture of how anionic phospholipids contribute so profoundly to the enzymatic activity of membrane-bound protease complexes involved in blood clotting (3).Membranes composed of mixed phospholipids (and other lipids) can form membrane microdomains with locally different surface properties. A noted example is the formation of cholesterol-and sphingolipid-rich lipid rafts and caveolae on cell surfaces (4, 5). Experiments using giant unilamellar vesicles have shown that even liposomes containing simple binary mixtures of neutral and anionic phospholipids spontaneously form anionic phospholipid-rich me...

The combination of profound muscle wasting and severe weight loss that occurs in heart failure is a complex phenomenon that involves the interplay of numerous factors. In this article, we describe processes that contribute to cachexia, as part of the clinical sequelae of heart failure, and their potential underlying mechanisms. While multiple mechanisms of cardiac cachexia have been described, we propose a multifactorial etiology for this condition that includes, but is not limited to, nutritional and gastrointestinal alterations, immunological and neurohormonal activation, and anabolic and catabolic imbalance.

Blood clotting reactions, such as those catalyzed by the tissue factor/factor VIIa complex (TF:VIIa), assemble on membrane surfaces containing anionic phospholipids such as phosphatidylserine (PS). In fact, membrane binding is critical for the function of most of the steps in the blood clotting cascade. In spite of this, our understanding of how the membrane contributes to catalysis, or even how these proteins interact with phospholipids, is incomplete. Making matters more complicated, membranes containing mixtures of PS and neutral phospholipids are known to spontaneously form PS-rich membrane microdomains in the presence of plasma concentrations of calcium ions, and it is likely that blood clotting proteases such as TF:VIIa partition into these PS-rich microdomains. Unfortunately, little is known about how membrane microdomain composition influences the activity of blood clotting proteases, which is typically not under experimental control even in "simple" model membranes. Our laboratories have developed and applied new technologies for studying membrane proteins to gain insights into how blood clotting protease-cofactor pairs assemble and function on membrane surfaces. This includes using a novel, nanoscale bilayer system (Nanodiscs) that permits assembling blood clotting protease-cofactor pairs on stable bilayers containing from 65 to 250 phospholipid molecules per leaflet. We have used this system to investigate how local (nanometerscale) changes in phospholipid bilayer composition modulate TF:VIIa activity. We have also used detailed molecular dynamics simulations of nanoscale bilayers to provide atomic-scale predictions of how the membrane-binding domain of factor VIIa interacts with PS in membranes.Most steps in the blood clotting cascade require the assembly of a serine protease together with its cognate regulatory protein on a membrane surface [1]. Both the coagulation serine protease (for example, factors VIIa, IXa or Xa) and its protein cofactor (tissue factor, factor VIIIa or factor Va, respectively) contain specific membrane-interactive domains that direct these proteins to assemble and function on membrane surfaces. Furthermore, the protein substrates (factors IX, X and prothrombin) of these membrane-bound enzymes also bind reversibly to membranes. Membrane surfaces can only support the binding and assembly of clotting factors if they contain exposed anionic phospholipids, with phosphatidylserine (PS) being the most active. But membrane binding does more than just localize these enzymes and substrates to regions of tissue trauma; the membrane also plays a critical role in catalysis, as blood clotting enzymes are thousands of times less active when released from the membrane surface. In spite of the critical importance of membrane binding, we still have a very incomplete understanding of how the membrane surface enhances blood clotting reactions. Furthermore, we lack a detailed understanding of how the membrane binding domains of blood clotting proteins actually interact with anionic phosph...

Since its emergence in December 2019, the virus known as severe acute respiratory syndrome coronavirus 2 has quickly caused a pandemic. This virus causes a disease now known as coronavirus disease 2019, or COVID-19. As an increasing proportion of the at-risk population becomes infected, and patients with severe illness are hospitalized, it is essential for hospitalists to remain current on how to best care for people with suspected or confirmed disease. Establishing a system for logistical planning, and accurate information sharing is strongly recommended. Infection control remains the ultimate goal. As such, health care workers should be educated on universal and isolation precautions, and the appropriate use of personal protective equipment. Social distancing should be encouraged to prevent the spread of infection, and creative and innovative ways to reduce contact may need to be considered. Moreover, it is imperative to prepare for contingencies as medical staff will inevitably get sick or become unavailable. Hospitalists have the difficult task of caring for patients while also adapting to the many logistical and social elements of a pandemic.

Summary. The clotting cascade requires the assembly of protease-cofactor complexes on membranes with exposed anionic phospholipids. Despite their importance, proteinmembrane interactions in clotting remain relatively poorly understood. Calcium ions are known to induce anionic phospholipids to cluster, and we propose that clotting proteins assemble preferentially on such anionic lipid-rich microdomains. Until recently, there was no way to control the partitioning of clotting proteins into or out of specific membrane microdomains, so experimenters only knew the average contributions of phospholipids to blood clotting. The development of nanoscale membrane bilayers (Nanodiscs) has now allowed us to probe, with nanometer resolution, how local variations in phospholipid composition regulate the activity of key protease-cofactor complexes in blood clotting. Furthermore, exciting new progress in solid-state NMR and large-scale molecular dynamics simulations allow structural insights into interactions between proteins and membrane surfaces with atomic resolution.