Membranes and Proteins A key di erence between prokaryo6c and eukaryo6c cells is in the elabora6on of internal membranes Membranes de ne organelles Each membrane bound compartment in a eukaryo5c cell has its own unique set of soluble proteins and its own unique set of membrane proteins A major focus in the next few lectures will be how this is accomplished Membranes perform several important cellular func6ons Separate one compartment from another membranes are selec5vely permeable barriers across which solutes are transported Provide a sca old for biochemical ac5vi5es one key example is energy transduc5on in mitochondria and chloroplasts Mediate some kinds of cell cell interac5ons Key element of many signal transduc5on pathways Membranes are uid lipid bilayers studded with proteins and oHen contain areas of di ering composi5on called ra s that oat around like icebergs on the ocean Three primary components of biological membranes Lipids These form the membrane bilayer itself Cholesterol a special type of lipid A ects bilayer proper5es Found only in eukaryotes Proteins These are associated with the bilayer or inserted into it to add func5on Three classes of membrane lipids Phospholipids Glycolipids and Sterols These are all amphipathic molecules Hydrophilic end and hydrophobic end 1 Phospholipids All have a phosphate linkage to a head group and 2 faRy acid chain tails Phosphoglycerides Major component of most membranes Consist of two faRy acids linked to glycerol with di ering chemical groups added to glycerol phosphate in the head group One faRy acid chain is saturated one unsaturated Many di erent types with di erent structures Sphingomyelin Sphingosine amino group instead of glycerol linkage to phosphate in head Two saturated faRy acid chains 2 Glycolipids Sphingosine amino group links to sugars instead of phosphate in head Have two saturated faRy acid chains 3 Sterols are amphipathic four ring hydrocarbons Cholesterol can increase or decrease membrane uidity depending on condi5ons Saturated faRy acid chains give rise to thicker and less uid bilayers Phospholipids Glycolipids Sterols Wikipedia Lipids in water can form two types of structures micelles and bilayers You need to get the hydrophobic tails of phospholipids away from water There are two common ways of accomplishing this Micelles Small sphere with tails pointed in Bilayers Two layers of lipids with tails pointed toward each other Which structure is formed is dependent on the type chemistry charge on head tail length tail shape and concentra5on of the lipid Biological phospholipids can form bilayers Two lea ets Proper6es of Lipid Bilayers Bilayers close upon themselves to make a con5nuous surface interac5ng with water No edges are leH exposed water interac5ng with hydrophobic tails unfavorable Membranes try to reseal if broken or punctured A cell will die if the seal does not reform fast enough Biological Membrane Composi6on Di erent cellular membranes are composed of di erent amounts of lipids and cholesterol and have very di erent protein composi5on The di erences in composi5on relate to di erences in func5on Each biological membrane has di erent proper5es based upon the di erent molecules used to make it Di erent membranes have di erent lipid composi6ons Lipid composi6on a ects bilayer thickness and membrane curvature Membranes are dynamic structures summary of movements of phospholipids transverse Using microscopy to determine the lateral mobility of lipids in the plane of the membrane Fluorescence recovery aHer photobleaching FRAP Label phospholipids with a uorescent probe Shine a bright laser on a small spot of membrane to bleach the uorescence on those lipids Measure how long it takes for other uorescent lipids to di use into the black region un5l it is as bright as the rest of the membrane ArfGAP1 dynamics and its role in COPI coat assembly on Golgi membranes of living cells Wei Liu1 Rainer Duden3 Robert D Phair2 and Jennifer LippincoR Schwartz1 Secretory protein tra cking relies on the COPI coat which by assembling into a laece on Golgi membranes concentrates cargo at speci c sites and deforms the membranes at these sites into coated buds and carriers The GTPase ac5va5ng protein GAP responsible for catalyzing Arf1 GTP hydrolysis is an important part of this system but the mechanism whereby ArfGAP is recruited to the coat its stability within the coat and its role in maintenance of the coat are unclear Here we use FRAP to monitor the membrane turnover of GFP tagged versions of ArfGAP1 Arf1 and coatomer in living cells ArfGAP1 underwent fast cytosol Golgi exchange with 40 of the exchange dependent on engagement of ArfGAP1 with coatomer and Arf1 and a ected by secretory cargo load Permanent ac5va5on of Arf1 resulted in ArfGAP1 being trapped on the Golgi in a coatomer dependent manner These data suggest that ArfGAP1 coatomer and Arf1 play interdependent roles in the assembly disassembly cycle of the COPI coat in vivo ArfGAP1 dynamics and its role in COPI coat assembly on Golgi membranes of living cells Wei Liu1 Rainer Duden3 Robert D Phair2 and Jennifer LippincoR Schwartz1 Secretory protein tra cking relies on the COPI coat which by assembling into a laece on Golgi membranes concentrates cargo at speci c sites and deforms the membranes at these sites into coated buds and carriers The GTPase ac5va5ng protein GAP responsible for catalyzing Arf1 GTP hydrolysis is an important part of this system but the mechanism whereby ArfGAP is recruited to the coat its stability within the coat and its role in maintenance of the coat are unclear Here we use FRAP to monitor the membrane turnover of GFP tagged versions of ArfGAP1 Arf1 and coatomer in living cells ArfGAP1 underwent fast cytosol Golgi exchange with 40 of the exchange dependent on engagement of ArfGAP1 with coatomer and Arf1 and a ected by secretory cargo load Permanent ac5va5on of Arf1 resulted in ArfGAP1 being trapped on the Golgi in a coatomer dependent manner These data suggest that ArfGAP1 coatomer and Arf1 play interdependent roles in the assembly disassembly cycle of the COPI coat in vivo Structure and Function of Biological Membranes MCB 5025 Spring semester 3 credits Instructor Alder Course description Overview of cell membrane structure and function based on a foundation of physical and biochemistry principles Topics include lipid structure and metabolism the lipid bilayer matrix membrane protein structure function and biogenesis membrane bioenergetics and