Trends and Prospects of Plant Proteases in Therapeutics.

The main function of proteases in any living organism is the cleavage of proteins resulting in the degradation of damaged, misfolded and potentially harmful proteins and therefore providing the cell with amino acids essential for the synthesis of new proteins. Besides this main function, proteases may play an important role as signal molecules and participate in numerous protein cascades to maintain the vital processes of an organism. Plant proteases are no exception to this rule.

Collagenolytic Enzymes and their Applications in Biomedicine.

Nowadays, enzymatic therapy is a very promising line of treatment for many different diseases. There is a group of disorders and conditions, caused by fibrotic and scar processes and associated with the excessive accumulation of collagen that needs to be catabolized to normalize the connective tissue content. The human body normally synthesizes special extracellular enzymes, matrix metalloproteases (MMPs) by itself.

Molecular mechanism of type I collagen homotrimer resistance to mammalian collagenases.

Type I collagen cleavage is crucial for tissue remodeling, but its homotrimeric isoform is resistant to all collagenases. The homotrimers occur in fetal tissues, fibrosis, and cancer, where their collagenase resistance may play an important physiological role. To understand the mechanism of this resistance, we studied interactions of alpha1(I)(3) homotrimers and normal alpha1(I)(2)alpha2(I) heterotrimers with fibroblast collagenase (MMP-1).

Structural heterogeneity of type I collagen triple helix and its role in osteogenesis imperfecta.

We investigated regions of different helical stability within human type I collagen and discussed their role in intermolecular interactions and osteogenesis imperfecta (OI). By differential scanning calorimetry and circular dichroism, we measured and mapped changes in the collagen melting temperature (DeltaTm) for 41 different Gly substitutions from 47 OI patients. In contrast to peptides, we found no correlations of DeltaTm with the identity of the substituting residue.

Selective retention and degradation of molecules with a single mutant alpha1(I) chain in the Brtl IV mouse model of OI.

We investigated the secretion, matrix incorporation and interactions of molecules with one and two mutant alpha1(I) collagen chains in the Brtl IV murine model for Osteogenesis Imperfecta, carrying a Gly-349 to Cys substitution in one col1a1 allele. We detected a significant deviation from the expected 25 and 50% content of the molecules with no (37-46%) and one (26-40%) mutant chains in skin and bone as well as in fibroblast and osteoblast cell culture media. Steady-state labeling with (35)S-Cys demonstrated incomplete secretion of the mutant collagen in cell culture, particularly molecules containing one mutant chain.

Prolyl 3-hydroxylase 1 deficiency causes a recessive metabolic bone disorder resembling lethal/severe osteogenesis imperfecta.

A recessive form of severe osteogenesis imperfecta that is not caused by mutations in type I collagen has long been suspected. Mutations in human CRTAP (cartilage-associated protein) causing recessive bone disease have been reported. CRTAP forms a complex with cyclophilin B and prolyl 3-hydroxylase 1, which is encoded by LEPRE1 and hydroxylates one residue in type I collagen, alpha1(I)Pro986.

Structure, stability and interactions of type I collagen with GLY349-CYS substitution in alpha 1(I) chain in a murine Osteogenesis Imperfecta model.

Here we report the structural and functional studies of collagen from the Brtl mouse, a heterozygous knock-in model for Osteogenesis Imperfecta, which has a G349C substitution introduced in one col1a1 allele. We observed that 25+/-5% of alpha 1(I) chains in different tissues and in different extracts from matrix deposited by cultured cells were S-S-linked mutant dimers. Apparently mutant and normal molecules are equally well incorporated into the matrix and they form mature covalent crosslinks with the same efficiency.

Changes in thermal stability and microunfolding pattern of collagen helix resulting from the loss of alpha2(I) chain in osteogenesis imperfecta murine.

Homozygous mutations resulting in formation of alpha1(I)(3) homotrimers instead of normal type I collagen cause mild to severe osteogenesis imperfecta (OI) in humans and mice. Limited studies of changes in thermal stability of type I homotrimers were reported previously, but the results were not fully consistent. We revisited this question in more detail using purified tendon collagen from wild-type (alpha1(I)(2)alpha2(I) heterotrimers) and oim (alpha1(I)(3)) mice as well as artificial alpha1(I)(3) homotrimers obtained by refolding of rat-tail-tendon collagen.