Human cathepsin D (CatD), a pepsin-family aspartic protease, plays an important role in tumor progression and metastasis. Here, we report the development of biomimetic inhibitors of CatD as novel tools for regulation of this therapeutic target. We designed a macrocyclic scaffold to mimic the spatial conformation of the minimal pseudo-dipeptide binding motif of pepstatin A, a microbial oligopeptide inhibitor, in the CatD active site. A library of more than 30 macrocyclic peptidomimetic inhibitors was employed for scaffold optimization, mapping of subsite interactions, and profiling of inhibitor selectivity. Furthermore, we solved high-resolution crystal structures of three macrocyclic inhibitors with low nanomolar or subnanomolar potency in complex with CatD and determined their binding mode using quantum chemical calculations. The study provides a new structural template and functional profile that can be exploited for design of potential chemotherapeutics that specifically inhibit CatD and related aspartic proteases.
- MeSH
- biomimetické materiály chemická syntéza chemie metabolismus toxicita MeSH
- Caco-2 buňky MeSH
- cyklické peptidy chemická syntéza chemie metabolismus toxicita MeSH
- enzymatické testy MeSH
- inhibitory proteas chemická syntéza chemie metabolismus toxicita MeSH
- kathepsin D antagonisté a inhibitory chemie metabolismus MeSH
- kinetika MeSH
- lidé MeSH
- molekulární struktura MeSH
- pepstatiny chemie MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Plant growth regulating properties of brevicompanines (Brvs), natural products of the fungus Penicillium brevicompactum, have been known for several years, but further investigations into the molecular mechanism of their bioactivity have not been performed. Following chemical synthesis of brevicompanine derivatives, we studied their activity in the model plant Arabidopsis by a combination of plant growth assays, transcriptional profiling, and numerous additional bioassays. These studies demonstrated that brevicompanines cause transcriptional misregulation of core components of the circadian clock, whereas other biological read-outs were not affected. Brevicompanines thus represent promising chemical tools for investigating the regulation of the plant circadian clock. In addition, our study also illustrates the potential of an unbiased -omics-based characterization of bioactive compounds for identifying the often cryptic modes of action of small molecules.
- MeSH
- Arabidopsis účinky léků fyziologie MeSH
- biologické přípravky chemická syntéza farmakologie MeSH
- cirkadiánní rytmus účinky léků MeSH
- cyklické peptidy chemická syntéza farmakologie MeSH
- fyziologie rostlin účinky léků MeSH
- genetická transkripce účinky léků MeSH
- indoly chemická syntéza farmakologie MeSH
- kořeny rostlin účinky léků růst a vývoj MeSH
- Penicillium chemie MeSH
- Publikační typ
- časopisecké články MeSH
In the venom of eusocial bee Lasioglossum laticeps, we identified a novel unique antimicrobial peptide named lasiocepsin consisting of 27 amino acid residues and two disulfide bridges. After identifying its primary structure, we synthesized lasiocepsin by solid-phase peptide synthesis using two different approaches for oxidative folding. The oxidative folding of fully deprotected linear peptide resulted in a mixture of three products differing in the pattern of disulfide bridges. Regioselective disulfide bond formation significantly improved the yield of desired product. The synthetic lasiocepsin possessed antimicrobial activity against both Gram-positive and -negative bacteria, antifungal activity against Candida albicans, and no hemolytic activity against human erythrocytes. We synthesized two lasiocepsin analogs cyclized through one native disulfide bridge in different positions and having the remaining two cysteines substituted by alanines. The analog cyclized through a Cys8-Cys25 disulfide bridge showed reduced antimicrobial activity compared to the native peptide while the second one (Cys17-Cys27) was almost inactive. Linear lasiocepsin having all four cysteine residues substituted by alanines or alkylated was also inactive. That was in contrast to the linear lasiocepsin with all four cysteine residues non-paired, which exhibited remarkable antimicrobial activity. The shortening of lasiocepsin by several amino acid residues either from the N- or C-terminal resulted in significant loss of antimicrobial activity. Study of Bacillus subtilis cells treated by lasiocepsin using transmission electron microscopy showed leakage of bacterial content mainly from the holes localized at the ends of the bacterial cells.
- MeSH
- antibakteriální látky chemická syntéza chemie farmakologie MeSH
- antifungální látky chemická syntéza chemie farmakologie MeSH
- Candida albicans účinky léků MeSH
- cyklické peptidy chemická syntéza chemie farmakologie MeSH
- cystin chemická syntéza chemie MeSH
- erytrocyty účinky léků MeSH
- gramnegativní bakterie účinky léků MeSH
- grampozitivní bakterie účinky léků ultrastruktura MeSH
- hemolýza MeSH
- lidé MeSH
- mikrobiální testy citlivosti MeSH
- molekulární sekvence - údaje MeSH
- sekundární struktura proteinů MeSH
- sekvence aminokyselin MeSH
- sekvenční analýza proteinů MeSH
- včelí jedy chemická syntéza chemie farmakologie MeSH
- včely chemie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Cyclic peptides 2a-2c, derived from the sequence of the C-terminal shortened analogs of the oostatic decapeptide H-Tyr-Asp-Pro-Ala-Pro-Pro-Pro-Pro-Pro-Pro-OH (1a), were synthesized and assayed on their effect in a reproduction of the flesh fly Neobellieria bullata. The cyclization of the N-terminal linear tetra- and pentapeptides 1b and 1c to the cyclotetra- and cyclopentapeptides 2b and 2c decreased the oostatic activity by one order of magnitude. The cyclodecapeptide 2a, which emerged spontaneously during the pentapeptide cyclization, was quite inactive. Comparative 1H and 13C NMR study on a conformation of the cyclopeptides 2a-2c, and their linear precursors 1b and 1c revealed that a space structure of the cyclic analogues 2b and 2c is too restricted to adopt a biological conformation necessary for receptor binding and therefore only minor oostatic activity is observed after their application. The lack of the oostatic activity in the case of the more flexible dimeric analogue 2a is ascribed to the size of its molecule and its overall shape that is not compatible with a receptor binding.
- MeSH
- biotest MeSH
- cyklické peptidy farmakologie chemická syntéza chemie MeSH
- Diptera fyziologie účinky léků MeSH
- financování organizované MeSH
- hmyzí proteiny chemie MeSH
- izotopy uhlíku MeSH
- magnetická rezonanční spektroskopie metody MeSH
- molekulární konformace MeSH
- molekulární modely MeSH
- oligopeptidy farmakologie chemická syntéza chemie MeSH
- oocyty fyziologie účinky léků MeSH
- protony MeSH
- zvířata MeSH
- Check Tag
- ženské pohlaví MeSH
- zvířata MeSH
