Cross-linking, collagen synthesis

Some proteins, like collagen and elastin, have covalent cross-links between their fibres, which are formed after their synthesis (post-translational modification). 

Synthetic peptides have been extensively used to study the thermal stability and folding of the triple helix. These peptides can be synthesized as either single chains or cross-linked peptides. Early on, such peptides were synthesized by polycondensation of tri- or hexapeptides, which led to a broad mass distribution that was difficult to separate. With the advances of solid-phase synthesis methods, peptides with defined chain length became available. The most studied collagen-like peptides are (Gly-Pro-Pro) or (Pro-Pro-Gly) and (Gly-Pro-4(if)Hyp) or (Pro-4(if)Hyp-Gly) with n varying from 5 to 15. Sutoh and Noda" " introduced the concept... 

Figure 3.4 Factors affecting foreign body reaction and potential points of intervention at the level of the myofibroblast (1) inhibit synthesis or release of TGF-P (2) block stimulation by TGF-P of its membrane receptors on the activated fibroblast (3) inhibit the Smad proteins, which transfer the TGF-P effect to the nucleus (4) inhibit transcription of procollagen mRNA (5) inhibit translation of the message to form procollagen (6) inhibit prolyl-4-hydroxylase, which creates hydroxyproline and facilitates helix formation (7) inhibit lysyl oxidase, which cross-links the collagen (8) enhance the function of MMPs, which degrade collagen, or inhibit TIMPs, which degrade MMPs.

It comes as no great surprise that trace elements may affect the growth and development of bone. Trace element deficiences profoundly alter hone metabolism in animals either directly or indirectly (3). The absence of a trace element in the diet can lead to inefficient functioning of a specific enzyme or enzymes that require the transition element as a cofactor. An example of this is the role of Cu and iron (Fe) in the cross-linking of collagen and elastins (4-9). The participation of Mn in the biosynthesis of mucopolysaccharides (10-12) is another example. Zn deficiency causes a reduction in osteoblastic activity, collagen and chondroitin sulfate synthesis and alkaline phosphatase activity (13-16). 

The copper metalloenzymes are involved in oxygen-using reactions. These enzymes include cytochrome c oxidase (respiratory chain), lysyl oxidase (collagen synthesis), and dopamine [3-hydroxylase (neurotransmitter synthesis). Lysyl oxidase is a small protein with a molecular weight of 32 kDa. This enzyme contains an unusual modification, namely cross-linking between two different parts of its polypeptide chain. The cross-linked region consists of a structure called lysine tyrosylquinone (Klinman, 1996). Two amino acids are involved in this cross-linked structure, and these are Lys 314 and Tyr 349. Lysine tyrosylquinone is used as a cofactor and is necessary for the catalytic activity of the enzyme. Other copper metalloenzymes contain a related cofactor, namely 2,4,5-tiihydrox5q5henylalanine (topaquinone, TPQ). Serum amino oxidase is a copper metalloenzyme that contains TPQ. TPQ consists of a modified residue of phenylalanine. The copper in the active site of the enzyme occurs immediately adjacent to the TPQ cofactor. 

The bone collagen cross-link (+)-deoxypyrrololine has potential clinical utility in the diagnosis of osteoporosis and other metabolic bone diseases. Intrigued by its novel structure and its promise to allow the early discovery of various bone diseases, the research team of M. Adamczyk developed a convergent total synthesis for this 1,3,4-trisubstituted pyrrole amino acid. The key step of the synthesis was the union of the nitroalkane and aldehyde fragments to obtain a diastereomeric mixture of the expected -nitro alcohol in good yield. This new functionality served as a handle to install the pyrrole ring. 

Adamczyk, M., Johnson, D. D., Reddy, R. E. Bone collagen cross-links a convergent synthesis of (+)-deoxypyrrololine. J. Org. Chem. 

Most signs of scurvy can be related to inadequate or abnormal collagen synthesis. Ascorbate enhances prolyl and lysyl hydroxylase activities (Chapter 25). Collagen formed in scorbutic patents is low in hydroxyproline and poorly cross-linked, resulting in skin lesions, bone fractures, and rupture of capillaries and other blood vessels. The absolute amount of collagen made in scorbutic animals may also decrease independently of the hydroxylation defect. The anemia of scurvy may result from a defect in iron absorption or folate metabolism. 

The synthesis of fibrillar collagen (e.g., types I, II, and III) begins Inside the cell with the chemical modification of newly made a chains and their assembly Into triple-helical procollagen within the endoplasmic reticulum. After secretion, procollagen molecules are cleaved, associate laterally, and are covalently cross-linked Into bundles called fibrils, which can form larger assemblies called fibers (see Figure 6-20). 

Tosylmethyl isocyanide (TosMIC) was employed in a key step for the synthesis of bicyclic heterocycles from L-glutamic acid <01H(55)2099>, while benzyl isocyanoacetate was utilized for reaction with a-acetoxynitro compound 41 to yield the pyrrole 42, an intermediate in the synthesis of (+)-deoxypyrro oline, a putative cross-link of bone collagen <01JOC11>. 

Boyera N, Galey I, Bernard BA. Effect of vitamin C and its deriwitives on collagen synthesis and cross-linking by normal human fibroblasts. IntJ Cosmet Sci 1998 20 151-8. 

Synthesis and maturation of interstitial collagens up to their final cross-linking into insoluble cross-banded fibrils is not the subject of this presentation, but it is necessary to mention some of the steps involved in the maturation of collagen molecules in order to understand the effects of vitamin C. 

Copper provides the essential functional part of a number of enzymes involved in oxidation and reduction reactions, including dopamine P-hydroxylase in the synthesis of noradrenaline and adrenaline, cytochrome oxidase in the electron transport chain (section 3.3.1.2) and superoxide dismutase, one of the enzymes involved in protection against oxygen radicals (section 7.4.3.1). Copper is also important in the oxidation of lysine to form the cross-links in collagen and elastin. In copper deficiency the bones are abnormally fragile, because the abnormal collagen does not permit the normal flexibility of the bone matrix. More importantly, elastin is less elastic than normal and copper deficiency can lead to death following rupture of the aorta. 

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