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The Typical Cell • typical cell: 1. nucleus 2. cell membrane 3. cytoplasm -cytosol -cytoskeleton 4. cytoplasmic organelles -membranous -non-membranous Cytoplasm • • • semi-fluid-like jelly within the cell division into two subdivisions: cytosol & organelles also contains a supportive framework of proteins: cytoskeleton Cytosol Cytosol -intracellular fluid of the cell -about 55% of the cell’s volume -about 70-90% water PLUS dissolved nutrients ions soluble & insoluble proteins waste products glucose, ATP amino acids, fatty acids •higher K+ •lower Na+ •higher concentration of dissolved and suspended proteins (enzymes, organelles) •lower concentration of carbohydrates (due to catabolism) •larger reserves of amino acids (anabolism) ECF •lower K+ •higher Na+ •lower concentration of dissolved and suspended proteins •higher concentration of carbohydrates •smaller reserves of amino acids Cytoskeleton: •internal framework of the cell •gives the cytoplasm flexibility and strength •three major components 1. microfilaments 2. intermediate filaments 3. microtubules microfilaments = made of thin filaments called actin -forms a dense network immediately under the PM + scattered throughout the cytoplasm (most prevalant at the cell periphery) -function: 1. anchor integral proteins and attaches them to the cytoplasm 2. interacts with larger filaments made up of myosin - results in active movements of the cell (e.g. muscle cells) or changes in cell shape -provide much of the mechanical strength of the cell + give the cell its shape -also provide support for cellular extensions called microvilli (small intestines) intermediate filaments = made up of vimentin, desmin or keratin -function: 1. impart strength 2. stabilize organelles 3. transport materials -some IFs are made of specialized proteins found in specific cell types e.g. neurofilaments in neurons - for transport of synaptic vesicles containing neurotransmitters •five major groups of IFs 1. Type I: acidic keratins (epithelial cells) 2. Type II: basic keratins (epithelial cells) 3. Type III: vimentin (bone, cartilage) -desmin (muscle) 4. Type IV: Neurofilaments (neuronal) 5. Type V: Lamins A, B, C (all cells) microtubules = repeating units of tubulin -assembly is controlled in the MTOC (microtubule organizing center) - located near the nucleus – region of tubulin proteins -for the assembly of tubulin into microtubules, centrioles and cilia -also known as a centrosome -role in MT assembly -found near the Golgi apparatus during interphase - called the centrosome (pair of centrioles at right angles) -MTs grow out from the centrosome -microtubule – a cylinder = 13 rows of tubulin arranged as a “straw” -straws can be glued together to form a doublet or a triplet -function: 1. cell shape & strength 2. organelles: anchor & movement 3. mitosis - form the spindle (chromosome movement) 4. form many of the non-membranous organelles (cilia, flagella, centrioles) Non-membranous Organelles A. Centrioles: short cylinders of tubulin – arranged as 9 short microtubule triplets -called a 9+0 array (9 peripheral triplets, 0 in the center) -each centriole is made of a pair – arranged perpendicular to one another -role in mitosis - spindle and chromosome alignment B. Cilia & Flagella •cilia = contain 9 groups of microtubule doublets surrounding a central pair -called a 9+2 array -anchored to a basal body just beneath the cell surface (9+0 array) -exposed portion of the cilia covered by PM – but the cilia itself is non-membranous -beat rhythmically to transport material -found in linings of several major organs covered with mucus -function in “cleaning” cilia 9+2 basal body 9+0 •flagella = resemble cilia -much larger -found singly -functions to move a cell through the ECF ??? What is the only human cell to possess a flagella??? Ribosomes = can be considered a nonmembranous organelle •2 protein subunits in combination with RNA -large 60S subunit = 28S rRNA (ribosomal RNA) + 50 proteins -small 40S subunit = 18S rRNA + 33 proteins •actual site of mRNA translation -> peptide strand •in association with the ER = where the peptide strand is fed into from the ribosome •also float freely within the cytoplasm as groups = polyribosomes Membranous Organelles •completely surrounded by a phospholipid bilayer similar to the PM surrounding the cell •allows for isolation of each individual organelle - so that the components of each organelle does not mix with the cytosol •therefore requires a well-coordinated system of transport for organelles to communicate and function together -”vesicular transport” - small transport vesicles pinch off one organelle, travel and then fuse to another organelle Overview of the secretory pathway 1. Endoplasmic reticulum (ER) = series of membrane-bound, flattened sacs in communication with the nucleus and the PM -each sac or layer = cisternae -inside or each sac = lumen -two types: Rough ER - outside studded with ribosomes -continuous with the nuclear membrane -protein synthesis, phospholipid synthesis -the peptide strand as it is being translated by the ribosome is fed into the RER -initial site of processing (attachment of carbohydrates) and sorting for transport to the Golgi -three functions: 1. synthesis 2. storage 3. transport • • • • • • • transport from the ribosome across the ER membrane requires the presence of a signal sequence 16-30 amino acids at the beginning of the peptide sequence (N-terminal) this signal sequence will vary from protein to protein by will have a few characteristics in common – starts with one or two positively charges amino acids and is followed by 6-12 hydrophobic AAs A complex of proteins will bind this signal in the cytoplasm = signal recognition particle/SRP the ER membrane has receptor for the SRP – docks the translating ribosome (yellow protein in figure) actual entrance of the protein requires the formation of a “hole” in the ER membrane (blue protein in figure) = translocon (series of multiple transmembrane proteins) this process also requires energy which is provided by GTP Translocation • this animation might be a bit complicated – but give it a try • http://www.rockefeller.edu/pubinfo/proteint arget.html Modifications in the RER • 1. formation of disulfide bonds – help stabilize the tertiary and quaternary structure of proteins • 2. folding of the peptide chain – misfolded proteins remain in the cytoplasm and are degraded – only properly folded proteins get transported to the Golgi for additional processing and transport – many proteins located in the ER which stimulate this folding • 3. addition and processing of carbohydrates • 4. breakage of specific peptide bonds – proteolytic cleavage • 5. assembly into multimeric proteins (more than one chain) for an animation go to http://sumanasinc.com/webcontent/animations/content/protei nsecretion_mb.html Smooth ER – extends from the RER but is free of ribosomes - many enzymes on the surface of the SER for: - 1. lipid and steroid (e.g. estrogen and testosterone) biosynthesis for membranes - 2. detoxification of toxins and drugs - 3. vesicle formation - 4. cleaves glucose so it can be released into the bloodstream -the amount of SER per cell can increase with drug use – cell accomadates to increase protection 3. Golgi Apparatus = stack of 3 to 20 flattened membrane sacs/cisternae •site of protein modification, and final packaging of the finished protein into secretory vesicles -> exocytosis or for use in the cytosol •movement of protein through the stacks via transport vesicles -definite direction: first stack = cis-face (from the RER, protein modification) middle stack = medial-face (adds carbohydrates) last stack = trans-face (modification and packaging into vesicles) Modifications in the Golgi • glycosylation = glycoprotein – most plasma membrane and secreted proteins have one or more carbohydrate chains that help target them to the correct location – some glycosylation occurs in the ER, others in the various sacs of the Golgi – in the Golgi are the addition of N- and O-linked oligosaccharides – specific sugar residues – O-linked are added one at a time in the Golgi to the amino acids serine, threonine or lysine (one to four saccharide subunits total) • added on by enzymes called glycosyltransferases • human A, B and O antigens are sugars added onto proteins and lipids inserted in the PM of the RBC • everyone has the glycosyltransferase needed to produce the O antigen – N-linked saccharides are linked together (about 14 sugars!) in the ER • transported out of the ER and imported into the Golgi • are attached specifically to asparagine amino acids within a protein • specific sugars can be removed or new ones added in the Golgi – changes the composition of the resulting glycoprotein – the sugars are found in the cytoplasm but are transported into the Golgi by specific transporters for their addition Modifications in the Golgi • some PM proteins and most secretory proteins are synthesized as larger, inactive proproteins that will require additional processing to become active – e.g. albumin, insulin, glucagon • this processing occurs very late in maturation = trans face – processing is catalyzed by protein-specific enzymes called proteases – some proteases are unique to the specific secretory protein – occurs in secretory vesicles that bud from the trans-Golgi face – processing could be at one site (albumin) others may require more than one peptide bond (insulin) -proteins processed in the Golgi will have unique sequences of amino acids that tell the protein where to go -these are called sorting signals -the lack of this signal means you will be secreted targets: 1. secretory vesicles for exocytosis 2. membrane vesicles for incorporation into PM 3. transport vesicles for intracellular destinations e.g. digestive enzymes to the lysosome -requires unique sorting signals 4. Mitochondria = site of energy production (ATP production) -via Cellular Respiration - breakdown of glucose into water and CO2 results in the production of ATP -initial glucose breakdown occurs in the cytosol -terminal stages occur in the mitochondria = Oxidative Phosphorylation -has its own DNA - maternal -reproduce themselves via dividing -consists of an outer mitochondrial membrane, an inner mitochondrial membrane and a fluid-filled space = mitochondrial matrix (contains ribosomes!) -the inner membrane is folded into folds called cristae (increase the membrane surface area for the enzymes of Oxidative Phosphorylation) -2 membrane layers: •outer - 50% lipid & 50% protein -very permeable - contains pores •inner - 20% lipid & 80% protein -less permeable -folded extensively to form partitions = cristae -contains proteins that transport H+ out of the lumen of the mitochondria -> electrochemical gradient -contains enzymes that use this gradient for the synthesis of ATP -also contains pumps to move ATP into the cytosol •matrix - lumen of the mitochondria -breakdown of glucose into water and CO2 ends here and results in the production of ATP Cellular respiration -glycolysis -citric acid cycle -electron transport chain http://biology.about.com/gi/dynamic/offsite.htm?site=http://www.sp.uconn.edu/%7Eterry/images/anim/ETS.html http://biology.about.com/gi/dynamic/offsite.htm?site=http://www.biocarta.com/pathfiles/krebPathway.asp http://vcell.ndsu.nodak.edu/animations/etc/movie.htm Glycolysis • • • • • • • literally means “splitting sugar” conversion of glucose (6 carbon sugar) into 2 molecules of pyruvate (3 carbon sugar) pyruvate will be converted into acetyl-coenzyme A (Acetyl-CoA) which then enters the citric acid cycle under aerobic conditions glucose is oxidized into pyruvate and then continues on to the citric acid cycle under anaerobic conditions glucose is oxidized into pyruvate then converted into lactate (lactic acid) reactions of glycolysis take place in the cytosol this pathway also creates the building blocks that are used for the synthesis of long chain fatty acids phosphorylation isomerization phosphorylation cleavage 2 ATP consumed no energy created key enzyme in controlling the rate of glycolysis = phosphofructokinase -liver enzyme that is inhibited when ATP levels are high http://web.indstate.edu/thcme/mwking/glycolysis.html http://science.nhmccd.edu/biol/glylysis/glylysis.html Glycolysis • we consume sugars other than glucose • two other abundant sugars are fructose and galactose • fructose can be converted into glyceraldehyde using the fructose-1phosphate pathway which utilizes different enzymes but still creates glyceraldehyde 3-phosphate • alternatively fructose can be phosphorylated to fructose-6phosphate by the same enzyme that phosphorylates glucose (hexokinase) – then is phosphorylated again = fructose-1,6phosphate • galactose can be converted into glucose-6-phosphate in a series of four steps using four different enzymes Glycolysis • the cells in all kinds of organisms convert glucose to pyruvate using similar reactions • however pyruvate can be processed in many ways to produce – 1. ethanol – by yeast • pyruvate – acetylaldehyde – ethanol • regulated by aldehyde dehydrogenase (DH = removes H+ from one substrate and adds it to another) • the opposite (alcohol to aldehyde) is catalyzed by alcohol dehydrogenase (ADH) – 2. lactate – by cells in the absence of oxygen – 3. acetyl CoA – the majority of pyruvate is processed by this next phase Citric Acid cycle • • • • • • • • • pyruvate is transported from the cytosol across the membranes of the mitochondria into its matrix interacts with coenzyme A (mitochondrial enzyme) to produce acetyl-CoA forms the waste product carbon dioxide (2 molecules) Acetyl-CoA is converted into oxaloacetic acid -> citric acid the citric acid is converted into a series of compounds that eventually regenerates OA acid takes place in the matrix and inner membrane of the mitochondria cycle results in the formation of NAD+ (nicotinamide adenine dinucleotide) – capable of storing high energy electrons as the cycle runs - NAD+ is reduced to form NADH and H+ (gains two electrons) NADH = electron carrier – – • • • NAD+ is capable of adding one H+ and 2 electrons GTP hydrolysis required. this NADH will then enter the electron transport chain Also generates the FADH2 electron carrier (for 4 electrons) while this cycle only runs in the presence of oxygen – no oxygen is used For each glucose this cycle has to “turn” Twice!! Electron transport chain • • • • • • • • protons are pumped from the matrix into the space between the inner and outer mitochondrial cytochrome c cytochrome c NADH membranes by enzyme complexes oxidase dehydrogenase reductase in addition, electrons are transferred from these complexes to oxygen eventually forming water and carbon dioxide protons are taken from NADH and pumped across the inner mitochondrial membrane by NADH dehydrogenase at the same time electrons are protons moved from NADH electrons dehydrogenase to cytochrome c reductase (by Q = ubiquinone) cytochrome c reductase pumps more protons (from water) across the membrane and more electrons are transferred to cytochrome c oxidase (by cytochrome c) cytochrome c oxidase also pumps more protons (taken from water) this creates a proton gradient = energy as protons flow down their gradient back into the matrix, they pass through an enzyme called ATP synthase – which synthesizes ATP http://biology.about.com/gi/dynamic/offsite.htm?site=http://www.sp.uconn.edu/%7Eterry/images/anim/ETS.html ETC animations • http://www.youtube.com/watch?v=xbJ0nbz t5Kw&feature=relmfu • http://www.youtube.com/watch?v=3y1dO4 nNaKY • http://biology.about.com/gi/dynamic/offsite. htm?site=http://www.sp.uconn.edu/%7Eter ry/images/anim/ETS.html 5. Lysosomes = “garbage disposals” -dismantle debris, eat foreign invaders/viruses taken in by endocytosis or phagocytosis -also destroy worn cellular parts from the cell itself and recycles the usable components = autophagy -form by budding off the Golgi -vesicle sacs that contain powerful enzymes to breakdown substances into their component parts e.g. nucleases = breakdown RNA & DNA into nucleotides proteases = breakdown proteins into amino acids -over 60 kinds of enzymes within the lysosome -these enzymes are collectively known as acid hydrolases -acidic interior - critical for function of these enzymes -created and maintained by a hydrogen pump that transport H+ into the interior - Active transport -chloride ions that diffuse in passively through a chloride channel - forms hydrochloric acid (HCl) 6. Peroxisomes: abundant in liver and kidney cells -main function is the breakdown of long chain fatty acids through their oxidation -oxidation is done by oxidases = enzymes that use oxygen to oxidize substances - such as peroxidase -oxidation reactions by these oxidases generate toxic oxidative chemicals such as hydrogen peroxide (H2O2) -therefore peroxisomes contain enzymes to break up these oxidative substances e.g. catalase to break this peroxide down - catalyze the breakdown of H2O2 into water and oxygen -outer membrane contains a variety of enzymes for 1. synthesis of bile acids 2. breakdown of lipids - oxidation of fatty acids and amino acids (formed during normal metabolic processes) 3. detoxification of alcohol – converts it into sugars F-actin and peroxisomes Diseases at the Organelle Level 1. Tay Sachs and lysosomes: lack one of the 40 lysosomal enzymes -produces a lysosomal storage disease -> build up of fatty material on nerve cells -failure of nervous system communication 2. Adrenoleukodystrophy and peroxisomes: peroxisomes lack an enzyme on the outer membrane which transports an essential enzyme into the peroxisome -leads to a build up of a long-chain fatty acid on cells of the brain and spinal cord -> loss of the myelin sheath -lethargy, skin darkens, blood sugar drops, altered heart rhythm imbalanced electrolytes, paralysis, death *** slowed by a certain triglyceride found in rapeseed oil Lorenzo Odone = “Lorenzo’s Oil”