scholarly journals Purification and characterization of the N-terminal propeptide of human type III procollagen

1985 ◽  
Vol 232 (1) ◽  
pp. 145-150 ◽  
Author(s):  
O Niemelä ◽  
L Risteli ◽  
J Parkkinen ◽  
J Risteli
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Terminal Segment ◽  
Purification And Characterization ◽  
Type Iii ◽  
Bacterial Collagenase ◽  
Type Iii Procollagen ◽  
Collagenous Domain ◽  
Bovine Type

The N-terminal propeptide of type III procollagen was purified from human ascitic fluid by using (NH4)2SO4 precipitation, DEAE-Sephacel chromatography at pH 8.6, Sephacryl S-300 chromatography and another DEAE-Sephacel chromatography at pH 4.5. The Mr of the human peptide was about 42 000, which corresponds in size to the propeptide released by the specific N-proteinase during the extracellular processing of collagen. Bacterial-collagenase digestion of the human peptide produced three fragments, which could be separated on a Bio-Gel P-10 column. The human propeptide and its collagenase-derived fragments, an N-terminal non-collagenous domain Col 1, a C-terminal non-helical domain Col 2 and a collagenous domain Col 3, resembled those derived from the N-terminal segment of bovine type III procollagen in their amino acid composition. The human peptide was found to contain sulphate, which may explain its extremely low isoelectric point (3.1). Antibodies against the human N-terminal propeptide reacted similarly with both the purified human peptide and a corresponding segment of bovine type III procollagen. The human propeptide could be used in developing radioimmunoassays for monitoring fibrotic processes.

scholarly journals Characterization of the Amino-Terminal Segment in Type III Procollagen

1976 ◽  
Vol 70 (1) ◽  
pp. 205-216 ◽  
Author(s):  
Hans NOWACK ◽  
Bjorn R. OLSEN ◽  
Rupert TIMPL
Keyword(s):  
Terminal Segment ◽  
Type Iii ◽  
Amino Terminal ◽  
Type Iii Procollagen

scholarly journals Improved purification and characterization of the OXA-2 β-lactamase

1984 ◽  
Vol 224 (3) ◽  
pp. 1009-1013 ◽  
Author(s):  
S Holland ◽  
J W Dale
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Terminal Region ◽  
Beta Lactamase ◽  
Purification And Characterization

An improved and scaled-up procedure has been developed for purifying the OXA-2 plasmid-mediated beta-lactamase. This has enabled us to improve the characterization of this enzyme, including a revised determination of its amino acid composition and the sequence of the N-terminal region of the protein.

Purification and characterization of porcine κ-casein

1984 ◽  
Vol 51 (2) ◽  
pp. 259-266 ◽  
Author(s):  
Jutta Cerning-Beroard ◽  
Claude Zevaco
Keyword(s):  
Amino Acid ◽  
Affinity Chromatography ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Sugar Composition ◽  
Purification And Characterization ◽  
Acetylneuraminic Acid ◽  
Porcine Milk

Summaryκ-casein from porcine milk was isolated and purified by affinity chromatography on thiopropyl Sepharose followed by hydroxyapatite chromatography. The amino acid composition of this protein is similar to that of bovine κ-casein. The sugar composition was established and sow κ-casein was found to be highly glycosylated. It contains 12 N-acetylneuraminic acid, 10 N-acetylgalactosamine, 2 N-acetylglucosamine and 22 galactose residues.

scholarly journals Purification and characterization of three forms of glutathione transferase from Proteus mirabilis

1988 ◽  
Vol 255 (3) ◽  
pp. 971-975 ◽  
Author(s):  
C Di Ilio ◽  
A Aceto ◽  
R Piccolomini ◽  
N Allocati ◽  
A Faraone ◽  
...  
Keyword(s):  
Amino Acid ◽  
Isoelectric Focusing ◽  
Proteus Mirabilis ◽  
Gel Electrophoresis ◽  
Glutathione Transferase ◽  
Total Activity ◽  
Purification And Characterization ◽  
Substrate Specificities ◽  
Structural Differences

Three forms of glutathione transferase (GST) with pI values of 6.0, 6.4 and 7.3 were isolated from Proteus mirabilis AF 2924 by glutathione-affinity chromatography followed by isoelectric focusing, and their structural, kinetic and immunological properties were investigated. Upon SDS/polyacrylamide-slab-gel electrophoresis, all forms proved to be composed of two subunits of identical (22,500) Mr. GST-6.0 and GST-6.4 together account for about 95% of the total activity, whereas GST-7.3 is present only in trace amounts. Extensive similarities have been found between GST-6.0 and GST-6.4. These include subunit molecular mass, amino acid composition, substrate specificities and immunological characteristics. GST-7.3 also cross-reacted (non-identity) with antisera raised against bacterial GST-6.0. None of the antisera raised against a number of human, rat and mouse GSTs cross-reacted with the bacterial enzymes, indicating major structural differences between them and the mammalian GSTs. This conclusion is further supported by c.d. spectra.

Gene Cloning, Nucleotide Sequencing, and Purification and Characterization of the Low-Specificityl-Threonine Aldolase from Pseudomonas sp. Strain NCIMB 10558

1998 ◽  
Vol 64 (2) ◽  
pp. 549-554 ◽  
Author(s):  
Ji-Quan Liu ◽  
Saeko Ito ◽  
Tohru Dairi ◽  
Nobuya Itoh ◽  
Michihiko Kataoka ◽  
...  
Keyword(s):  
Amino Acid ◽  
Significant Loss ◽  
Site Directed Mutagenesis ◽  
Nucleotide Sequencing ◽  
Predicted Amino Acid Sequence ◽  
Amino Acid Residues ◽  
Purification And Characterization ◽  
Reading Frame ◽  
Threonine Aldolase

ABSTRACT A low-specificity l-threonine aldolase (l-TA) gene from Pseudomonas sp. strain NCIMB 10558 was cloned and sequenced. The gene contains an open reading frame consisting of 1,041 nucleotides corresponding to 346 amino acid residues. The gene was overexpressed in Escherichia colicells, and the recombinant enzyme was purified and characterized. The enzyme, requiring pyridoxal 5′-phosphate as a coenzyme, is strictlyl specific at the α position, whereas it cannot distinguish between threo and erythro forms at the β position. In addition to threonine, the enzyme also acts on various other l-β-hydroxy-α-amino acids, includingl-β-3,4-dihydroxyphenylserine,l-β-3,4-methylenedioxyphenylserine, andl-β-phenylserine. The predicted amino acid sequence displayed less than 20% identity with those of low-specificityl-TA from Saccharomyces cerevisiae,l-allo-threonine aldolase from Aeromonas jandaei, and four relevant hypothetical proteins from other microorganisms. However, lysine 207 of low-specificity l-TA from Pseudomonas sp. strain NCIMB 10558 was found to be completely conserved in these proteins. Site-directed mutagenesis experiments showed that substitution of Lys207 with Ala or Arg resulted in a significant loss of enzyme activity, with the corresponding disappearance of the absorption maximum at 420 nm. Thus, Lys207 of thel-TA probably functions as an essential catalytic residue, forming an internal Schiff base with the pyridoxal 5′-phosphate of the enzyme to catalyze the reversible aldol reaction.

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