Department of Medical Biochemistry Semmelweis University
Subcellular biochemistry
February-March 2017 Subcellular biochemistry (biochemical aspects of cell biology)
Miklós Csala
Semmelweis University Dept. of Medical Chemistry, Molecular Biology and Pathobiochemistry TOPICS
• COMPARTMENTATION
Basic structural and functional principle of the eukaryotic cell organization
• MEMBRANES
Plasma membrane, intracellular membranes, nucleus
• MOVEMENT AND TRAFFIC OF ORGANELLES
Cytoskeleton, microfilaments, microtubuli, actomyosin, vesicular transport
• METABOLISM AND TRANSPORT
Principles of metabolome, metabolic profile of various organelles COMPARTMENTATION
Basic structural and functional principle of the eukaryotic cell organization
Cellular membranes
The endomembrane system • nuclear envelope • endoplasmic reticulum (ER) • Golgi apparatus • lysosome (phagosome, autophagosome) • endosome • transport vesicles • cell membrane • peroxisome (?)
Mitochondria • inner and outer mitochondrial membranes
Biological membranes: the fluid mosaic model Biological membranes: Amphipathic membrane lipids Biological membranes: Amphipathic membrane lipids Biological membranes: Amphipathic membrane lipids Biological membranes: Amphipathic membrane lipids Biological membranes: Amphipathic membrane lipids Biological membranes: Amphipathic membrane lipids Biological membranes: Lipid bilayer Biological membranes: Integral and peripheral membrane proteins Associated membrane proteins With transmembrane helices or β-barrels Integral membrane proteins With lipid anchors
Glycophosphatidylinositol (GPI) anchor Carbohydrate layer around the cell membrane Roles of cellular membranes • Separating cell contents from the outside environment (plasma membrane). • Cell recognition and signaling. • Creating separated intracellular compartments (endomembranes and mitochondrial membranes). • Holding the components of some metabolic pathways in place (compartment or membrane). • Regulating the transport of materials into or out of cells.
Roles of membrane proteins Mitochondrial respiratory chain: a central metabolic process based entirely on compartmentation and membrane structure Compartment
• membrane-encompassed • characteristic proteome • characteristic metabolome • characteristic pH, redox and ion millieu • separation and connection • driving cellular specialization and development of tissues and organs Cellular compartments
• cytosol
Formed by the endomembrane system • nucleus • endoplasmic reticulum (ER) • endoplasmic reticulum - Golgi intermediate compartment (ERGIC) • Golgi apparatus • lysosome (phagosome, autophagosome) • endosome (early, late, MVT) • transport vesicles • peroxisome (?) • lipid droplet • autophagic vacuole
Formed by mitochondrial membranes • mitochondrial matrix • mitochondrial intermembrane space
Plasma membrane Plasma membrane: Membrane microdomains: lipid rafts
Cholesterol- and sphingolipid-enriched, highly dynamic, submicroscopic (25–100 nm diameter) assemblies, which float in the liquid- disordered lipid bilayer in cell membranes. Plasma membrane: Caveola - invaginated lipid rafts
Roles in signal transduction and endocytosis Plasma membrane: Asymmetry
Phosphatidylcholine and glycolipids
Phosphatidylserine and phosphatidylinositols Plasma membrane: Asymmetry
ATP ATP
Apoptosis: „eat me” signal Platelet factor 3
Lipid raft organization and endocytosis
Signaling protein recruitment The endomembrane system The endoplasmic reticulum
The origin of the endomembrane system
Function protein synthesis (RER) lipid synthesis biotransformation Ca2+-storage glucose production The dinamic endoplasmic reticulum ER subdomains and the specialized ER
• Rough ER (RER) • Smooth ER (SER) • Transitional ER (tER) / ER exit site (ERES) • Junctional ER (jER) / sub-plasmalemmal ER / plasma membrane associated membrane (PAM) • Mitochondrion-associated membrane (MAM) • Sarcoplasmic reticulum (SR) The (ER-directing) signal sequence Attachment at the translocon channel Ribosome cycles and SRP cycle Signal peptidase N-glycosylation in the ER Golgi-appartus
Distribution and Shipping Department
Function glycoprotein maturation O-glycosylation limited proteolysis protein sorting synthesis of sphingomyelin and glycolipids synthesis of proteoglycans, polysaccharides and glucosaminoglycans Lysosome Its enzymes can break up almost anything. • Acid hydrolases – DNAse, RNAse – proteases (e.g. cathepsins) – glycosidases – lipases – phosphatases – sulfatases etc. • V-ATPase (vesicular type proton ATPase) • Fusion with late endosome or autophagosome • Lipofuscin „wear and tear” pigment granules accumulating in aging cells around the nucleus Autophagosome
Degrading the cell's own components using the lysosomal machinery Endosomes and lysosomes Recicling endosome
Lysosome
Early endosome
Multivesicular body LDL-cholesterol uptake Peroxysome Function oxidative catabolism with low energy conservation certain processes of lipid synthesis (ether-phospholipid, cholesterol, bile acids) hydrogen peroxide production and elimination
Crystalloid core Lipid droplet Nucleus
The control center of the cell
DNA (nuclear genome) Histones and non-histone proteins Transcription factors RNA synthesis and processing Synthesis of ribosomes (nucleolus) Regulation of gene expression Nucleus
Double bi-layer
Nuclear lamina (lamin protein)
Nuclear pores across two membranes
Outer membrane continuous with the RER (ribosomes on both)
Perinuclear space continuous with the RER lumen The nuclear pore The Ran GTPase cycle Nuclear import / export with Ran Ran-GTP makes the import receptor unloaded or the export receptor loaded Evolution of endomembranes and formation of mitochondria by endosymbiosis Mitochondria
Power Plants of the Cell
Function Oxidative phosphorylation ATP synthesis (energy conservation) driven by oxidative catabolism of fuel molecules
Oxidative catabolism of fatty acids, pyruvate and amino acids (β- oxidation, PDH and citrate cycle)
Urea cycle, hem synthesis
Ca2+ buffering and storage
Heat production Mitochondria
Matrix nucleoid, ribosomes, large enzyme complexes
Inner membrane impermeable protein:lipid ratio > 3:1 - transporters for metabolites and proteins Outer membrane - enzymes of oxidative phosphorylation pores (porin) - proteins of fusion and fission permeable for up to 5 KDa Mitochondrial genome
mtDNA: • circular, double stranded • heavy (G) and light (C) strands • 100-10000 copy/cell • replicated by DNApolγ • 37 genes • no introns • little noncoding sequences • unique code (mitochondrial tRNA) Mitochondrial genome
13 proteins 22 tRNAs 2 rRNAs Comparison between human nuclear and mitochondrial genomes Mitochondrial diseases
Unique inheritance Mitochondrial encephalomyopathy, lactic acidosis, and stroke
Motor neuron diseases Myoclonic epilepsy with ragged red fibers Mitochondrial genome genetic relationships of individuals or groups within a species, phylogeny among different species