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Chapter 8 Fibrous proteins Major fibrous protein of epithelial tissues is keratin Major fibrous proteins of connective tissue are: Collagen Elastin Dr. Stephen C. Hardies7-3735437D hardies@uthscsa.edu Slide 2 The fundamental building block of a keratin fiber is a coiled coil dimer of a type I and type II keratin polypeptide. A. coiled coil B. C. heptad repeat: hxxhxxx, where "h" means hydrophobic and "x" means any residue. -helix -helical coiled coil domain Slide 3 -Keratin Where: Major protein of hair, skin, nails, some oral mucosa; small amounts in all epithelial cells. Cellular location: Intracellular; not found in connective tissues Structure: fibrous bundle of coil-coiled -helixes. Crosslinks: disulfide bonds Multigene family: family members differ by: tissue structureHardSoft amt. CysMoreLess amt. CrosslinksMoreLess Function: Provide insoluble structural protein for body surfaces Slide 4 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a) Proteins are linear chains of amino acids with polarity. Polarity means that there is directionality caused by adjacent amino acids being joined from COOH group to NH 2 group. One end of a polypeptide will be termed the N- terminus and the other the C-terminus. b)Disulfide bonds are crosslinks between sulfur atoms of either cysteine residues or methionine residues. c) An alpha helix can accommidate any sequence of amino acids. Slide 5 Collagen Where: 30% of total protein. Major protein of connective tissues: bones, tendons, ligaments, basement membranes, dentin, cementum, (not enamel). Cellular location: extracellular matrix. Structure: triple helix (tropocollagen). Subsequent to secretion, tropocollagen is assembled and crosslinked to make insoluble collagen fibers. Function: Provides tensile strength to soft connective tissues. Tissues that must be elastic but exhibit tensile strength (e.g. ligaments) have a mixture of collagen and elastin. Collagen fibers in bone reinforce against fracture. Slide 6 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a) In woven bone, collagen fibers are laid down in a disorganized array, whereas in lamellar bone they are laid down in a more organized parallel fashion. b) Woven bone is stronger than lamellar bone. c) Laminar bone is made by remodeling woven bone. Slide 7 Features of collagen primary structure -Gly-Pro-Met-Gly-Pro-Ser-Gly-Pro-Arg- -Gly-Leu-Hyp-Gly-Pro-Hyp-Gly-Ala-Hyp- -Gly-Pro-Gln-Gly-Phe-Gln-Gly-Pro-Hyp- Repetitive character of sequence: Slide 8 Collagen 3 stranded helix Slide 9 The collagen triple helix is stabilized by an interchain hydrogen bonding network involving the hydroxyl group of hydroxyproline, the glycine carbonyl group, and water molecules. Slide 10 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a)Hydroxyproline and hydroxylysine are among the 20 amino acids inserted into polypeptides by ribosomes. b)Glycine is NH 2 -CH 2 -COOH. c)Glycine is the only amino acid that doesnt have a D and an L isomer. Slide 11 Type I collagen fibril EM 670 angstrom D period tropocollagen Key assembly interaction Gap Slide 12 gap Ca 10 (OH) 2 (PO 4 ) 6 Slide 13 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a)Ca 10 (OH) 2 (PO 4 ) 6 is the same mineral (hydroxyapatite) that forms enamel. b)Whereas bone is reinforced by collagen fibers, enamel uses a different protein named amelogenin. c)Enamel has a higher mineral content than bone. Slide 14 Slide 15 fibril 50 nm 5 nm Type VI Anchoring fiber Slide 16 Type IV collagen forms planar arrays and makes basement membranes Type VII collagen anchors basement membrane to underlying connective tissue (stromal) cell layer. Slide 17 Type III collagen Sometimes called elastic collagen or extensible collagen Slide 18 Collagen type III micrograph visualized in polarized light, showing crimped organization of the fibrils. Slide 19 Osteogenesis Imperfecta Blue sclera Dentinogenesis Imperfecta Opalescent and cracked teeth Defects in Type I collagen cause: Type I: associated with OI. Will probably need full crown coverage. Slide 20 Abraded teeth in Dentinogenesis Imperfecta Slide 21 Chronic bone fractures in Osteogenesis Imperfecta Slide 22 An example of a mutation underlying Osteogenesis Imperfecta 988 Pro Gly Pro Arg Gly Arg Thr Gly Asp Ala CCG GGT CCT CGC GGT CGC ACT GGT GAT GCT Pro Cys Pro Arg Gly Arg Thr Gly Asp Ala CCT TGT CCT CGC GGT CGC ACT GGT GAT GCT Slide 23 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a)DNA is a polymer of bases named A,T,G, and C. b)The sequence of bases in a gene determines the sequence of the polypeptide that will be produced. c)A mutation is a heritable change to the sequence of a gene that causes the encoded polypeptide to have altered function. d)Some individual residue changes may have large effects on the protein function, whereas other changes may have little or no effect. e)Sickle cell anemia is an example of an inherited disease caused by a single residue change in a protein. Slide 24 Ehlers-Danlos Syndrome Hyperplasticity of the skin Slide 25 Slide 26 Steps in collagen biosynthesis: Translation on rough ER and entry into ER. Hydroxylation in the ER. Triple helix assembly. Glycosylation, transport to Golgi, further glycosylation. Secretion. Removal of propeptides. Assembly into fibrils. Crosslinking. Slide 27 Collagen hydroxylation Slide 28 Slide 29 Perifollicular abnormalities. Gingival abnormalities Scurvy Slide 30 Slide 31 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl What are preliminary steps to get procollagen polypeptides into the ER? a)Depending on the synthetic cell type, transcription factors will select specific preprocollagen genes from a family of such genes be transcribed and translated. b)The preprocollagen polypeptide is released into the cytoplasm. c)A special sequence on the N-terminus called the signal peptide directs the preprocollagen to a pore in the ER through which it enters the ER. Slide 32 Examples of collagen nomenclature: An individual polypeptide: 1 (I) procollagen Assembled triple helix with propeptides still on: Type I procollagen Triple helix after propeptide removal: Type I tropocollagen Assembled into 50 nm fiber: Collagen microfibril Slide 33 Slide 34 A third lysine (from a third tropocollagen) can add to make pyridinoline. Crosslinks involving hydroxylysine are more stable than those involving lysine. There is also a crosslink involving histidine. Slide 35 Steps in collagen biosynthesis: Translation on rough ER and entry into ER. Hydroxylation in the ER. Triple helix assembly. Glycosylation, transport to Golgi, further glycosylation. Secretion. Removal of propeptides. Assembly into fibrils. Crosslinking. + (or else degrade) vitamin C + HOlys HOpro + + Slide 36 Elastin: Where: elastic connective tissues. Cellular location: extracellular matrix Function: add elasticity to connective tissue Structure: beta spiral Crosslink: desmosine Slide 37 Facial features associated with Williams Syndrome Dental features include small widely spaced teeth and malocclusion. Genetic defects in elastin underlie Williams Syndrome Slide 38 Dynamic structure of elastin: The structure of elastin is called a spiral, and is loosely held together by the hydrophobic force. It is easily deformed to an extended configuration, but will relax back to a compact conformation. Repeating unit: PGVGV Slide 39 Slide 40 Fibrillin Where: connective tissue; extracellular matrix Structure: forms outer envelope of elastin microfibrils Genetic defects in fibrillin result in Marfans Syndrome, characterized by a tall gaunt appearance, joint problems, an often leading to death by aortic aneurysm. Slide 41 Abraham Lincoln is thought to have had Marfans syndrome. Slide 42 Aortic Aneurysm Slide 43 Enzymes that turn over connective tissue Common names: collagenase, gelatinase, elastase Formal names: Matrix Metalloproteases (MMPs) Function: degrade connective tissue in support of tissue remodeling, wound healing, and cell migration (including during metastasis). Family of Zn ++ -requiring zymogens embedded in connective tissue. They can be activated in a cascade starting from a cell surface MMP. Slide 44 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl Zymogens are inactive forms of enzymes that will become activated by cleavage to remove a propeptide. Zymogenes are also called proenzymes. Examples of zymogens are: a)trypsinogen b)collagen c)coagulation factors d)DNA polymerase e)cathepsin Slide 45 Proteoglycan aggregate Where: major component of ground substance, the material within which collagen and other fibers are assembled to form connective tissue. Function: make space, absorb water and allow compressibility through water flow, act as reservoir of Ca ++ prior to mineralization. Slide 46 Proteoglycan aggregate Slide 47 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl Which of these are glycosaminoglycans? a) b) c) d) Slide 48 Proteoglycan breakdown associated with mineralization Slide 49 Adhesion proteins Where: cell surface Functions: adhere cells to extracellular proteins (or to other cells), sense presence of extracellular proteins, sense mechanical stress in tissues. Slide 50 Integrins cell membrane inactive Outside in signaling Inside out activation Slide 51 True or False? http://biochem.uthscsa.edu/hardies-bin/survey.pl a) Integrins would be a good choice of adhesion protein to use for a cell like a platelet that has to very rapidly switch from a nonbinding to a tight binding mode. Integrins bind the sequence RGD in ligand proteins. Weve seen that in: b) bone proteins c) matrix metalloproteases d) keratin Slide 52 Tissues of Teeth