Acidic cleavage of the aspartyl-proline bond and the limitations of the reaction

Jáuregui-Adell, J.; Martí, J.

doi:10.1016/0003-2697(75)90148-7

Cited by 72 publications

(31 citation statements)

References 8 publications

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“…Thus, the gene fusion construct encodes KSI-D-P-D-E-novispirin G10. The DP dipeptide is prone to mild acid hydrolysis at elevated temperatures (7,13), and the glutamic acid residue located immediately upstream of novispirin G10 is a substrate of a glutamyl endopeptidase I.…”

Section: Methodsmentioning

confidence: 99%

Effects of Intratracheal Administration of Novispirin G10 on a Rat Model of Mucoid Pseudomonas aeruginosa Lung Infection

Song

Mygind

et al. 2005

Antimicrob Agents Chemother

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Chronic Pseudomonas aeruginosa lung infection is a major problem for patients with cystic fibrosis (CF). The biofilm mode of growth of the pathogen makes it highly resistant to antibiotic treatment, and this is especially pronounced with mucoid strains. In this study, novispirin G10, a synthetic antimicrobial peptide patterned loosely on sheep myeloid antimicrobial peptide 29, was tested in a rat model of mucoid P. aeruginosa lung infection. P. aeruginosa NH57388A, a mucoid strain isolated from a CF patient, was mixed with the alginate produced by the bacterium itself and adjusted to a concentration of 10 10 CFU/ml. Each rat received 10 9 CFU of bacteria intratracheally in the left lung to establish lung infection. At 0 and 3 h post P. aeruginosa infection, the treated group of rats received novispirin G10 (0.1 mg/ml, 0.1 ml/rat) intratracheally, whereas the control group received vehicle treatment only. The animals were sacrificed on days 3, 5, 7, and 10 after challenge for evaluation of various parameters. On day 5, 50% of the rats in the treated group had cleared the bacteria from the lungs, whereas in the control group, none of the rats cleared the pathogen (P < 0.03). The average bacterial loads remaining in the lungs of treated rats on days 3 and 5 were more than 170-and 330-fold lower than in the control groups (P < 0.0005 and P < 0.0003). In accordance, the macroscopic and microscopic lung pathology was also significantly milder in the treated group compared to the control group (P < 0.0002). Lung cytokine responses in the treated group were significantly lower than in the control group. The results suggest that novispirin G10 might be useful in treating antibiotic-resistant P. aeruginosa lung infections.

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Section: Methodsmentioning

confidence: 99%

Effects of Intratracheal Administration of Novispirin G10 on a Rat Model of Mucoid Pseudomonas aeruginosa Lung Infection

Song

Mygind

et al. 2005

Antimicrob Agents Chemother

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“…Another sample of the amidinotransferase was cleaved in 7 M guanidine. HCl dissolved in 75% formic acid at 37°C for 60 h [16]. The reaction mixture was then diluted and soluble and insoluble fragments separated as above.…”

Section: Isolation Of Peptides and Sequence Analysismentioning

confidence: 99%

The amino acid sequences of human and pig l‐arginine:glycine amidinotransferase

1994

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We have isolated and sequenced the L-arginine:glycine amidinotransferase of pig kidney mitochondria. Due to endogenous proteolysis, the purified molecules showed some heterogeneity at the N terminus. The longest form recovered had 386 amino acids. Part of the pig amidinotransferase sequence information was used to isolate cDNA clones coding for the human enzyme. The deduced amino acid of the human amidinotransferase was 37 amino acids longer due to the presence of a signal sequence. The mature proteins were 94% identical to each other and 36% identical to the sequences of bacterial L-arginine:inosamine phosphate amidinotransferases.

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“…For carboxyl-terminal analysis, digestion was accomplished with 1 nmol of NAF and 0.1 unit of carboxypeptidase A, 0.01 unit of carboxypeptidase B, or 0.1 unit of carboxypeptidase Y (all from Sigma) according to established methods (11). Cleavage of the Asp-Pro bond of unmodified NAF (1 nmol) was performed in 50 ,ul of 75% (vol/vol) formic acid at 37°C for 48 hr (12 13.5, and centrifugation yielded a 516-g cell pellet.…”

Section: Methodsmentioning

confidence: 99%

Synthesis and expression in Escherichia coli of the gene encoding monocyte-derived neutrophil-activating factor: biological equivalence between natural and recombinant neutrophil-activating factor.

Lindley

Aschauer

Seifert

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

212

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The neutrophil-activating factor (NAF) purified from the conditioned medium of lipopolysaccharidestimulated human monocytes was sequenced and found to consist of 72 amino acids: SAKELRCQCIKTYSKPFHPKFI-KELRVIESGPHCANTEIIVKLSDGRELCLDP-KENWVQRVVEKFLKRAENS. Purified preparations of natural NAF contained, in addition to this main form, minor amounts of three amino-terminal variants with 77 (+AVLPR), 70, and 69 residues. A gene coding for the 72-amino acid NAF was synthesized, cloned, and expressed in Escherichia coli. Western (immunologic) blot analysis of crude bacterial extracts, with an antiserum raised against natural NAF, revealed a single band that comigrated with natural NAF. Recombinant NAF purified to homogeneity had identical amino-and carboxyl-terminal sequences to the 72-amino acid natural NAF. Recombinant NAF was tested on human neutrophils and had the same activity and potency as natural NAF in inducing chemotaxis, rapidly increasing cytosolic free Ca2+, activating the respiratory burst, and releasing specific and azurophilic granular contents.

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Acidic cleavage of the aspartyl-proline bond and the limitations of the reaction

Cited by 72 publications

References 8 publications

Effects of Intratracheal Administration of Novispirin G10 on a Rat Model of Mucoid Pseudomonas aeruginosa Lung Infection

Effects of Intratracheal Administration of Novispirin G10 on a Rat Model of Mucoid Pseudomonas aeruginosa Lung Infection

The amino acid sequences of human and pig l‐arginine:glycine amidinotransferase

Synthesis and expression in Escherichia coli of the gene encoding monocyte-derived neutrophil-activating factor: biological equivalence between natural and recombinant neutrophil-activating factor.

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