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Hydrolysis Activity (hydrolysis + activity)

Distribution by Scientific Domains

Life Sciences	40%

Selected Abstracts

Non-enzymatic hydrolysis of fluorescein diacetate (FDA) in a Mediterranean oak (Quercus ilex L.) litter

EUROPEAN JOURNAL OF SOIL SCIENCE, Issue 2 2008
E. Alarcón-Gutiérrez
Summary We show the presence of interfering substances when the total microbial activity in litter samples is measured with fluorescein diacetate (FDA), and we propose some methodological modifications to avoid such interference. Three distinct litter layers (the OhLn, the OhLv and the OhLf) of evergreen oak (Quercus ilex L.) were characterized by 13C CPMAS NMR and the spectra show that the recalcitrant aromatic and phenolic compounds increase with the degree of degradation of litter. A wide range of sources of interference in the hydrolysis of FDA was found. To understand the origin of this interference, sterilized litter materials (i.e. ,-irradiated or autoclaved) and a wide range of organic substances (i.e. amino acids, glucose, sorbitol and organic humic acids) were investigated. Insignificant differences on the FDA hydrolysis activity (FDA activity) were found in the ,-irradiated and non-irradiated OhLn litter, indicating that ,-irradiation does not destroy enzymes. Conversely, after heat-sterilization of litter, samples showed FDA activity corresponding to 60, 34.8 and 30.8% (in the OhLn, the OhLv and the OhLf layers, respectively) of that of control litters. This indicates the presence of non-enzymatic interfering substances in the FDA assays. As the humification and litter depth increased, hydrolysis of FDA due to interferences decreased, indicating degradation and/or chelation of interfering substances. We hypothesize that lysine, arginine, histidine and cysteine are mainly responsible for the hydrolysis of FDA. We suggest that the use of phosphate buffer (50 mm, pH 7.0) with incubation <,30 minutes, in combination with a temperature between 30 and 40°C, produces insignificant interference in the determination of the final FDA activity in litter samples. [source]

DLC-1 suppresses non-small cell lung cancer growth and invasion by RhoGAP-dependent and independent mechanisms

MOLECULAR CARCINOGENESIS, Issue 5 2008
Kevin D. Healy
Abstract Expression of the tumor suppressor deleted in liver cancer-1 (DLC-1) is lost in non-small cell lung (NSCLC) and other human carcinomas, and ectopic DLC-1 expression dramatically reduces proliferation and tumorigenicity. DLC-1 is a multi-domain protein that includes a Rho GTPase activating protein (RhoGAP) domain which has been hypothesized to be the basis of its tumor suppressive actions. To address the importance of the RhoGAP function of DLC-1 in tumor suppression, we performed biochemical and biological studies evaluating DLC-1 in NSCLC. Full-length DLC-1 exhibited strong GAP activity for RhoA as well as RhoB and RhoC, but only very limited activity for Cdc42 in vitro. In contrast, the isolated RhoGAP domain showed 5- to 20-fold enhanced activity for RhoA, RhoB, RhoC, and Cdc42. DLC-1 protein expression was absent in six of nine NSCLC cell lines. Restoration of DLC-1 expression in DLC-1-deficient NSCLC cell lines reduced RhoA activity, and experiments with a RhoA biosensor demonstrated that DLC-1 dramatically reduces RhoA activity at the leading edge of cellular protrusions. Furthermore, DLC-1 expression in NSCLC cell lines impaired both anchorage-dependent and -independent growth, as well as invasion in vitro. Surprisingly, we found that the anti-tumor activity of DLC-1 was due to both RhoGAP-dependent and -independent activities. Unlike the rat homologue p122RhoGAP, DLC-1 was not capable of activating the phospholipid hydrolysis activity of phospholipase C-,1. Combined, these studies provide information on the mechanism of DLC-1 function and regulation, and further support the role of DLC-1 tumor suppression in NSCLC. © 2007 Wiley-Liss, Inc. [source]

Communication between E,54, promoter DNA and the conserved threonine residue in the GAFTGA motif of the PspF ,54 -dependent activator during transcription activation

MOLECULAR MICROBIOLOGY, Issue 2 2004
Patricia Bordes
Summary Conversion of E,54 closed promoter complexes to open promoter complexes requires specialized activators which are members of the AAA (ATPases Associated with various cellular Activities) protein family. The ATP binding and hydrolysis activity of E,54 activators is used in an energy coupling reaction to remodel the E,54 closed promoter complex and to overcome the ,54 -imposed block on open complex formation. The remodelling target for the AAA activator within the E,54 closed complex includes a complex interface contributed to by Region I of ,54, core RNA polymerase and a promoter DNA fork junction structure, comprising the E,54 regulatory centre. One ,54 binding surface on E,54 activators is a conserved sequence known as the GAFTGA motif. Here, we present a detailed characterization of the interaction between Region I of ,54 and the Escherichia coli AAA ,54 activator Phage shock protein F. Using E,54 promoter complexes that mimic different conformations adopted by the DNA during open complex formation, we investigated the contribution of the conserved threonine residue in the GAFTGA motif to transcription activation. Our results suggest that the organization of the E,54 regulatory centre, and in particular the conformation adopted by the ,54 Region I and the DNA fork junction structure during open complex formation, is communicated to the AAA activator via the conserved T residue of the GAFTGA motif. [source]

Biochemical characterization of rab proteins from Bombyx mori,

ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 2 2009
Tomohide Uno
Abstract The small GTPases known as Rab proteins are key regulators of membrane trafficking. We used RT-PCR to isolate cDNA clones of insect-specific Rab proteins (BRabN1 and BRabN2) showing low homology with known Rab proteins from other animals, from mRNA of Bombyx mori. These 2 Rabs were produced in Escherichia coli and purified. BRabN1 bound [3H]-GDP and [35S]-GTP,S with dissociation constants of 0.087 × 10,6,M and 1.02 × 10,6,M, respectively, whereas those of BRabN2 were 0.546 × 10,6,M and 1.02 × 10,6,M, respectively. Binding of [35S]-GTP,S to BRabN1 and N2 was inhibited by GDP and GTP. The GTP-hydrolysis activities of BRabN1 and N2 were 154 and 35.5,mmol/min/mole, respectively, and bound [35S]-GTP,S was exchanged efficiently with GTP. BRabN1 also showed ATPase activity and exchange of [35S]-GTP,S with ATP. Monoclonal antibodies against BRabN1 and N2 did not recognize any other Rab proteins, and Western blotting using the anti-BRabN1 antibody revealed a single band in the testis of B. mori. These results suggest that BRabN1 and N2 of B. mori bind GTP, convert from the GTP-bound state to the GDP-bound state by intrinsic GTP hydrolysis activity, and return to the GTP-bound state with the exchange, and that BRabN1 is specifically expressed in testis. Arch. Insect Biochem. Physiol. 2008. © 2008 Wiley-Liss, Inc. [source]

Nitrile hydrolysis activity of Rhodococcus erythropolis NCIMB 11540 whole cells

BIOTECHNOLOGY JOURNAL, Issue 5 2006
Mandy K. S Vink
Abstract The nitrile hydrolyzing properties of the bacterium strain Rhodococcus erythropolis NCIMB 11540 have been investigated. Using whole cells of the microorganism, a wide variety of aromatic and aliphatic cyanide-containing substrates was successfully hydrolyzed to the corresponding amide or acid. In the case of dicyanides, selective monohydrolysis took place, which was further explored in the desymmetrization of malononitriles resulting in the corresponding cyano amides in enantiomeric excesses of up to 98%. [source]