Characterization of post-translational modifications in full-length human BMP-1 confirms the presence of a rare vicinal disulfide linkage in the catalytic domain and highlights novel features of the EGF domain
“…Interestingly, most of the non-identified sequence stretches of HRNR contain at least one cysteine residue (Fig. 2), which might have been caused either by a reduced detectability of Cys-containing peptides in LC-MS/MS analysis 13 or have been masked by the presence of posttranslational modifications, e.g. at Cys residues.Figure 1 P. a .-inhibitory antimicrobials in stratum corneum.…”
In the search for potential mechanisms underlying the remarkable resistance of healthy skin against infection by soil bacteria like
Pseudomonas
(
P
.)
aeruginosa
we identified fragments of the intrinsically disordered protein hornerin as potent microbicidal agents in the stratum corneum. We found that, independent of the amino acid (AA)-sequence, any tested linear cationic peptide containing a high percentage of disorder-promoting AA and a low percentage of order-promoting AA is a potent microbicidal antimicrobial. We further show that the antimicrobial activity of these cationic intrinsically disordered antimicrobial peptides (CIDAMPs) depends on the peptide chain length, its net charge, lipidation and environmental conditions. The ubiquitous presence of latent CIDAMP sources in nature suggests a common and yet overlooked adapted innate disinfection system of body surfaces. The simple structure and virtually any imaginable sequence or composition of disorder-promoting AA allow the generation of a plethora of CIDAMPs. These are potential novel microbicidal anti-infectives for various bacterial pathogens, including
P. aeruginosa
, methicillin-resistant
Staphylococcus aureus
(MRSA) and fungal pathogens like
Candida albicans
and
Cryptococcus neoformans
.
“…Interestingly, most of the non-identified sequence stretches of HRNR contain at least one cysteine residue (Fig. 2), which might have been caused either by a reduced detectability of Cys-containing peptides in LC-MS/MS analysis 13 or have been masked by the presence of posttranslational modifications, e.g. at Cys residues.Figure 1 P. a .-inhibitory antimicrobials in stratum corneum.…”
In the search for potential mechanisms underlying the remarkable resistance of healthy skin against infection by soil bacteria like
Pseudomonas
(
P
.)
aeruginosa
we identified fragments of the intrinsically disordered protein hornerin as potent microbicidal agents in the stratum corneum. We found that, independent of the amino acid (AA)-sequence, any tested linear cationic peptide containing a high percentage of disorder-promoting AA and a low percentage of order-promoting AA is a potent microbicidal antimicrobial. We further show that the antimicrobial activity of these cationic intrinsically disordered antimicrobial peptides (CIDAMPs) depends on the peptide chain length, its net charge, lipidation and environmental conditions. The ubiquitous presence of latent CIDAMP sources in nature suggests a common and yet overlooked adapted innate disinfection system of body surfaces. The simple structure and virtually any imaginable sequence or composition of disorder-promoting AA allow the generation of a plethora of CIDAMPs. These are potential novel microbicidal anti-infectives for various bacterial pathogens, including
P. aeruginosa
, methicillin-resistant
Staphylococcus aureus
(MRSA) and fungal pathogens like
Candida albicans
and
Cryptococcus neoformans
.
“…The only astacin proteinases resistant to fetuin-B inhibition were the members of the BMP-1/tolloid subfamily. As shown by crystal structure analysis 56 and mass spectrometry 57 , these enzymes have in common a unique vicinal disulfide bond located in the substrate binding β-strand in intimate distance to the zinc ion of the active site. This arrangement most likely prevents access of fetuin-B to the active site.…”
Vertebrate fetuins are multi-domain plasma-proteins of the cystatin-superfamily. Human fetuin-A is also known as AHSG, α2-Heremans-Schmid-glycoprotein. Gene-knockout in mice identified fetuin-A as essential for calcified-matrix-metabolism and bone-mineralization. Fetuin-B deficient mice, on the other hand, are female infertile due to zona pellucida ‘hardening’ caused by the metalloproteinase ovastacin in unfertilized oocytes. In wildtype mice fetuin-B inhibits the activity of ovastacin thus maintaining oocytes fertilizable. Here we asked, if fetuins affect further proteases as might be expected from their evolutionary relation to single-domain-cystatins, known as proteinase-inhibitors. We show that fetuin-A is not an inhibitor of any tested protease. In stark contrast, the closely related fetuin-B selectively inhibits astacin-metalloproteinases such as meprins and ovastacin, but not astacins of the tolloid-subfamily, nor any other proteinase. The analysis of fetuin-B expressed in various mammalian cell types, insect cells, and truncated fish-fetuin expressed in bacteria, showed that the cystatin-like domains alone are necessary and sufficient for inhibition. This report highlights fetuin-B as a specific antagonist of ovastacin and meprin-metalloproteinases. Control of ovastacin was shown to be indispensable for female fertility. Meprin inhibition, on the other hand, renders fetuin-B a potential key-player in proteolytic networks controlling angiogenesis, immune-defense, extracellular-matrix-assembly and general cell-signaling, with implications for inflammation, fibrosis, neurodegenerative disorders and cancer.
“…Recombinant human BMP-1 and Xenopus sizzled were produced and purified as previously described (40,72). Lawler and co-workers (73) with minor modifications.…”
Bone morphogenetic protein 1 (BMP-1) is an important metalloproteinase that synchronizes growth factor activation with extracellular matrix assembly during morphogenesis and tissue repair. The mechanisms by which BMP-1 exerts these effects are highly context dependent. Because BMP-1 overexpression induces marked phenotypic changes in two human cell lines (HT1080 and 293-EBNA cells), we investigated how BMP-1 simultaneously affects cell-matrix interactions and growth factor activity in these cells. Increasing BMP-1 led to a loss of cell adhesion that depended on the matricellular glycoprotein thrombospondin-1 (TSP-1). BMP-1 cleaved TSP-1 between the VWFC/procollagen-like domain and the type 1 repeats that mediate several key TSP-1 functions. This cleavage induced the release of TSP-1 C-terminal domains from the extracellular matrix and abolished its previously described multisite cooperative interactions with heparan sulfate proteoglycans and CD36 on HT1080 cells. In addition, BMP-1–dependent proteolysis potentiated the TSP-1–mediated activation of latent transforming growth factor–β (TGF-β), leading to increased signaling through the canonical SMAD pathway. In primary human corneal stromal cells (keratocytes), endogenous BMP-1 cleaved TSP-1, and the addition of exogenous BMP-1 enhanced cleavage, but this had no substantial effect on cell adhesion. Instead, processed TSP-1 promoted the differentiation of keratocytes into myofibroblasts and stimulated production of the myofibroblast marker α-SMA, consistent with the presence of processed TSP-1 in human corneal scars. Our results indicate that BMP-1 can both trigger the disruption of cell adhesion and stimulate TGF-β signaling in TSP-1–rich microenvironments, which has important potential consequences for wound healing and tumor progression.
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