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C-terminal Side (c-terminal + side)
Selected AbstractsATP allosteric activation of atrial natriuretic factor receptor guanylate cyclaseFEBS JOURNAL, Issue 11 2010Teresa Duda Atrial natriuretic factor receptor guanylate cyclase (ANF-RGC) is the receptor and the signal transducer of two natriuretic peptide hormones: atrial natriuretic factor and brain natriuretic peptide. It is a single transmembrane-spanning protein. It binds these hormones at its extracellular domain and activates its intracellular catalytic domain. This results in the accelerated production of cyclic GMP, a second messenger in controlling blood pressure, cardiac vasculature and fluid secretion. ATP is obligatory for the transduction of this hormonal signal. Two models of ATP action have been proposed. In Model 1, it is a direct allosteric transducer. It binds to the defined regulatory domain (ATP-regulated module) juxtaposed to the C-terminal side of the transmembrane domain of ANF-RGC, induces a cascade of temporal and spatial changes and activates the catalytic module residing at the C-terminus of the cyclase. In Model 2, before ATP can exhibit its allosteric effect, ANF-RGC must first be phosphorylated by an as yet unidentified protein kinase. This initial step is obligatory in atrial natriuretic factor signaling of ANF-RGC. Until now, none of these models has been directly validated because it has not been possible to segregate the allosteric and the phosphorylation effects of ATP in ANF-RGC activation. The present study accomplishes this aim through a novel probe, staurosporine. This unequivocally validates Model 1 and settles the over two-decade long debate on the role of ATP in ANF-RGC signaling. In addition, the present study demonstrates that the mechanisms of allosteric modification of ANF-RGC by staurosporine and adenylyl-imidodiphosphate, a nonhydrolyzable analog of ATP, are almost (or totally) identical. [source] Analysis of a dextran-binding domain of the dextranase of Streptococcus mutansLETTERS IN APPLIED MICROBIOLOGY, Issue 3 2002H. Morisaki Aims: To examine the dextran-binding domain of the dextranase (Dex) of Streptococcus mutans. Methods and Results: Deletion mutants of the Dex gene of Strep. mutans were prepared by polymerase chain reaction and expressed in Escherichia coli cells. Binding of the truncated Dexs to dextran was measured with a Sephadex G-150 gel. Although the Dexs which lacked the N-terminal variable region lost enzyme activity, they still retained dextran-binding ability. In addition, further deletion into the conserved region from the N-terminal did not influence the dextran-binding ability. However, the Dex which carried a deletion in the C-terminus still possessed both enzyme activity and dextran-binding ability. Further deletion into the conserved region from the C-terminal resulted in complete disappearance of both enzyme and dextran-binding activities. Conclusions: Deletion analysis of the Dex gene of Strep. mutans showed that the C-terminal side (about 120 amino acid residues) of the conserved region of the Dex was essential for dextran-binding ability. Significance and Impact of the Study: The dextran-binding domain was present in a different area from the catalytic site in the conserved region of the Dex molecule. The amino acid sequence of the dextran-binding domain of the Dex differed from those of glucan-binding regions of other glucan-binding proteins reported. [source] Atmospheric pressure matrix-assisted laser desorption/ionization ion trap mass spectrometry of sulfonic acid derivatized tryptic peptidesRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 23 2001T. Keough Atmospheric pressure matrix-assisted laser desorption/ionization (AP-MALDI) and ion trap mass spectrometry have been used to study the fragmentation behavior of native peptides and peptide derivatives prepared for de novo sequencing applications. Sulfonic acid derivatized peptides were observed to fragment more extensively and up to 28 times more efficiently than the corresponding native peptides. Tandem mass spectra of native peptides containing aspartic or glutamic acids are dominated by cleavage on the C-terminal side of the acidic residues. This significantly limits the amount of sequence information that can be derived from those compounds. The MS/MS spectra of native tryptic peptides containing oxidized Met residues show extensive loss of CH3SOH and little sequence-specific fragmentation. On the other hand, the tandem mass spectra of derivatized peptides containing Asp, Glu and oxidized Met show much more uniform fragmentation along the peptide backbone. The AP-MALDI tandem mass spectra of some derivatized peptides were shown to be qualitatively very similar to the corresponding vacuum MALDI postsource decay mass spectra, which were obtained on a reflector time-of-flight instrument. However, the ion trap mass spectrometer offers several advantages for peptide sequencing relative to current reflector time-of-flight instruments including improved product ion mass measurement accuracy, improved precursor ion selection and MSn. These latter capabilities were demonstrated with solution digests of model proteins and with in-gel digests of 2D-gel separated proteins. Copyright © 2001 John Wiley & Sons, Ltd. [source] Ortho-aminobenzoic acid-labeled bradykinins in interaction with lipid vesicles: Fluorescence studyBIOPOLYMERS, Issue 5 2002R. F. Turchiello Abstract The peptide hormone bradykinin (BK) (Arg1 -Pro2 -Pro3 -Gly4 -Phe5 -Ser6 -Pro7 -Phe8 -Arg9) and its shorter homolog BK1,5 (Arg1 -Pro2 -Pro3 -Gly4 -Phe5) were labeled with the extrinsic fluorescent probe ortho -aminobenzoic acid (Abz) bound to the N-terminal and amidated in the C-terminal carboxyl group (Abz-BK-NH2 and Abz-BK1,5 -NH2). The fragment des-Arg9 -BK was synthesized with the Abz fluorescent probe attached to the 3-amino group of 2,3-amino propionic acid (DAP), which positioned the Abz group at the C-terminal side of BK sequence, constituting the peptide des-Arg9 -BK-DAP(Abz)-NH2. The spectral characteristics of the probe were similar in the three peptides, and their fluorescent properties were monitored to study the interaction of the peptides with anionic vesicles of dimyristoylphosphatidylglycerol (DMPG). Time-resolved fluorescence experiments showed that the fluorescence decay of the peptides was best described by double-exponential kinetics, with mean lifetimes values around 8.0 ns in buffer pH 7.4 that increased about 10% in the presence of DMPG vesicles. About a 10-fold increase, compared with the values in aqueous solution, was observed in the steady-state anisotropy in the presence of vesicles. A similar increase was also observed for the rotational correlation times obtained from time-resolved anisotropy decay profiles, and related to the overall tumbling of the peptides. Equilibrium binding constants for the peptide,lipid interaction were examined monitoring anisotropy values in titration experiments and the electrostatic effects were evaluated through Gouy,Chapman potential calculations. Without corrections for electrostatic effects, the labeled fragment Abz-BK1,5 -NH2 presented the major affinity for DMPG vesicles. Corrections for the changes in peptide concentration due to electrostatic interactions suggested higher affinity of the BK fragments to the hydrophobic phase of the bilayer. © 2002 Wiley Periodicals, Inc. Biopolymers 65: 336,346, 2002 [source] | ||||||||||