Biochemistry: Molecular cloaking devices

Nature (Impact Factor: 41.46). 07/2008; 453(7197):861-2. DOI: 10.1038/453861a
Source: PubMed


Protease enzymes cut other proteins into pieces, but some can be blocked by inhibitors. One such inhibitor binds to the substrate rather than the enzyme, suggesting a new tactic for drug discovery.

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    • "Dominant-negative inhibition by co-expression of its non-functional mutant forms is a widely used strategy to inhibit the function of a protein (Tsiavaliaris et al., 2002; Wu et al., 2004; Overall and Blobel, 2007), whereby the mutants bind to the enzyme substrate without cleaving it, thereby limiting substrate availability for the wild-type enzyme. Alternatively the mutant could form a non-functional complex with the enzyme with high affinity and specificity, thus blocking the interaction between the wildtype enzyme and the substrate (Kodadek, 2008 ). This mutant provided a powerful tool for the functional studies described here. "
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    ABSTRACT: BACKGROUND Controlled trophoblast invasion into the maternal decidua (interstitial invasion) is important for placental development. Factors secreted by the maternal decidual cells and the extravillous trophoblast can influence trophoblast invasion and abnormalities in the invasion process may lead to pregnancy complications. Serine protease HtrA3 is highly expressed in the decidual cells in the late secretory phase of the menstrual cycle and throughout pregnancy, and in most trophoblast cell types, apart from the invading interstitial trophoblast during the first trimester. HtrA3 and its family members are down-regulated in a number of cancers and are proposed as tumour suppressors. The current study aimed to investigate whether HtrA3 is secreted by decidual cells, and whether inhibiting such secretion alters trophoblast invasion. METHODS AND RESULTS Human endometrial stromal cells (HESCs) were decidualized with estradiol, medroxyprogesterone acetate and cyclic adenosine monophosphate. Real-time RT–PCR, western blotting and immunocytochemistry confirmed that HtrA3 mRNA and protein expression increased during decidualization. HtrA3 was also detected in the conditioned media (CM) of the decidualized HESCs, confirming its secretion. For functional studies, a protease-inactive mutant form of HtrA3 which was previously confirmed to be a dominant-negative inhibitor was produced using wheat germ cell-free technology. CM from decidualized HESCs significantly suppressed invasion of trophoblast HTR-8 cells (P < 0.01), whereas inhibition of HtrA3 in this CM by exogenous HtrA3 mutant resulted in increased trophoblast HTR-8 cell invasion (P < 0.001). CONCLUSIONS These results strongly support the hypothesis that decidual HtrA3 negatively regulates trophoblast invasion.
    Preview · Article · Feb 2011 · Human Reproduction
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    • "Over-manifestation of the cellular protease is the kernel factor for genesis of various human body disorders. The development mechanism of a protease and their proteolysis of specific peptide (or protein) as substrate have played a pivotal role on genesis of inflammatory disease and outbreaks of cancer and their metastasis by sudden change of physiological condition and immune system [1]. Recently, synthetic polymers conjugated to drug (or specific molecules) via peptide linker chain have been employed as a targeted drug carrier [2]–[4]. "
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    ABSTRACT: Characterization and control of proteolysis of peptides by specific cellular protease is a priori requisite for effective drug discovery. Here, we report the nanomechanical, in situ monitoring of proteolysis of peptide chain attributed to protease (Cathepsin B) by using a resonant nanomechanical microcantilever immersed in a liquid. Specifically, the detection is based on measurement of resonant frequency shift arising from proteolysis of peptides (leading to decrease of cantilever's overall mass, and consequently, increases in the resonance). It is shown that resonant microcantilever enables the quantification of proteolysis efficacy with respect to protease concentration. Remarkably, the nanomechanical, in situ monitoring of proteolysis allows us to gain insight into the kinetics of proteolysis of peptides, which is well depicted by Langmuir kinetic model. This implies that nanomechanical biosensor enables the characterization of specific cellular protease such as its kinetics.
    Full-text · Article · Feb 2009 · PLoS ONE

  • No preview · Article · Aug 2008 · Nature Reviews Drug Discovery
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