Rapid Purification of Human DNase I Using Mouse Monoclonal Anti-DNase I Antibodies and Characterization of the Antibodies

2000 ◽  
Vol 17 (2) ◽  
pp. 71-76 ◽  
Author(s):  
Tamiko Nakajima ◽  
Toshihiro Yasuda ◽  
Haruo Takeshita ◽  
Yoshimitsu Nakashima ◽  
Shinjiro Mori ◽  
...  
Keyword(s):  
Dnase I ◽  
Rapid Purification

scholarly journals Differences in the interaction of the catalytic groups of the active centres of actinidin and papain Rapid purification of fully active actinidin by covalent chromatography and characterization of its active centre by use of two-protonic-state reactivity probes

1981 ◽  
Vol 197 (3) ◽  
pp. 739-746 ◽  
Author(s):  
K Brocklehurst ◽  
B S Baines ◽  
J P Malthouse
Keyword(s):  
Cysteine Proteinase ◽  
Thiol Group ◽  
Interactive System ◽  
Active Centre ◽  
Ph Values ◽  
Hydrophobic Binding ◽  
Rapid Purification ◽  
Active Centres ◽  
Covalent Chromatography

1. A rapid method of isolation of fully active actinidin, the cysteine proteinase from Actinidia chinensis (Chinese gooseberry or kiwifruit), by covalent chromatography, was devised. 2. The active centre of actinidin was investigated by using n-propyl 2-pyridyl disulphide, 4-(N-aminoethyl 2′-pyridyl disulphide)-7-nitrobenzo-2-oxa-1,3-diazole and 4-chloro-7-nitrobenzofurazan as reactivity probes. 3. The presence in actinidin in weakly acidic media of an interactive system containing a nucleophilic sulphur atom was demonstrated. 4. The pKa values (3.1 and 9.6) that characterize this interactive system are more widely separated than those that characterize the interactive active centre systems of ficin (EC 3.4.22.3) and papain (EC 3.4.22.2) (3.8 and 8.6, and 3.9 and 8.8 respectively). 5. Actinidin was shown to resemble ficin rather than papain in (i) the disposition of the active-centre imidazole group with respect to hydrophobic binding areas, and (ii) the inability of the active-centre aspartic acid carboxy group to influence the reactivity of the active-centre thiol group at pH values of about 4. 6. The implications of the results for one-state and two-state mechanisms for cysteine-proteinase catalysis are discussed.

Characterization of the rat intestinal Fc receptor (FcRn) promoter: transcriptional regulation of FcRn gene by the Sp family of transcription factors

2004 ◽  
Vol 286 (6) ◽  
pp. G922-G931 ◽  
Author(s):  
Lingling Jiang ◽  
Jiafang Wang ◽  
R. Sergio Solorzano-Vargas ◽  
Hugh V. Tsai ◽  
Edgar M Gutierrez ◽  
...  
Keyword(s):  
Transcriptional Regulation ◽  
Cell Lines ◽  
Promoter Region ◽  
Core Promoter ◽  
Dnase I ◽  
Fc Receptor ◽  
Minimal Promoter ◽  
Luciferase Reporter ◽  
Core Promoter Region

The regulatory elements that control the transcriptional regulation of the intestinal Fc receptor ( FcRn) have not been elucidated. The objective of this study was to characterize the core promoter region of the rat FcRn gene. Chimeric clones that contained various regions of the promoter located upstream of the luciferase reporter were transiently transfected into either IEC-6 or Caco-2 cell lines and nuclear extracts were used to perform DNase I footprint and DNA binding assays (EMSA). Transfection of chimeric upstream nested deletions-luciferase reporter clones into either of these cell lines supported robust reporter activity and identified the location of the minimal promoter at −157/+135. DNase I footprint analysis revealed two complexes located within the gene's core promoter region, and site-directed mutagenesis identified two regions that were critical to maintain basal expression. EMSA identified the presence of five Sp elements within the immediate promoter region that are capable of binding members of the Sp family of proteins. Among the five Sp elements, one element appears to not bind Sp1, Sp2, or Sp3 while influencing the interaction of Sp proteins with an adjacent Sp site. Overexpression of either Sp1 or Sp3 augments activity of the minimal promoter in Sp-deficient Drosophila SL2 cells. In summary, we report on the characterization of the rat FcRn minimal promoter, including the characterization of five Sp elements within this region that interact with members of the Sp family of transcriptional factors and drive promoter activity in intestinal cell lines.

Purification and Characterization of Subtilisin Cleaved Actin Lacking the Segment of Residues 43-47 in the DNase I Binding Loop

Biochemistry ◽  
1995 ◽  
Vol 34 (45) ◽  
pp. 14834-14842 ◽  
Author(s):  
Peter Kiessling ◽  
Werner Jahn ◽  
Gernot Maier ◽  
Bernhard Polzar ◽  
Hans Georg Mannherz
Keyword(s):  
Dnase I ◽  
Purification And Characterization ◽  
Binding Loop

Intein-mediated rapid purification and characterization of a novel recombinant agonist for VPAC2

Peptides ◽  
2006 ◽  
Vol 27 (6) ◽  
pp. 1359-1366 ◽  
Author(s):  
Rong-jie Yu ◽  
Qiu-ling Xie ◽  
Yun Dai ◽  
Yuan Gao ◽  
Tian-hong Zhou ◽  
...  
Keyword(s):  
Purification And Characterization ◽  
Rapid Purification
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