Purification and Characterization of a Dipeptidyl Carboxypeptidase from <i>Pseudomonas</i> sp. WO24

Ogasawara, Wataru; Abe, Nobuhide; Hagio, Takashi; Okada, Hirofumi; Morikawa, Yasushi

doi:10.1271/bbb.61.858

Cited by 9 publications

(5 citation statements)

References 11 publications

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“…The serine, cysteine, and aspartic protease inhibitors (PMSF, NBS, and WRK) had almost no effect, whereas the group selective modification regents DEPC (for histidine residues), EAM (for lysine residues), and CHD (for arginine residues) reduced the activity of His 6 -EcDCP to between 6 and 66.9% of its maximum. In agreement with other studies of microbial DCPs (Henrich et al 1993;Miyoshi et al 1992;Nagamori et al 1990;Ogasawara et al 1997), His 6 -EcDCP exhibited a strong susceptibility toward the chelating agents EDTA and 1,10-phenanthroline. Together with the presence of a potential Zn 2?…”

Section: Effects Of Metal Ions and Other Compounds On His 6 -Ecdcp Acsupporting

confidence: 91%

“…(Henrich et al 1993); PsDCP, Pseudomonas sp. DCP (Ogasawara et al 1997); PmDCP, Pseudomonas maltophilia DCP (Dasarathy et al 1989); BpDCP, Bacillus pumilus (Nagamori et al 1991); NvDCP, Neanthes virens DCP (Kawamura et al 2000); HsACE, Homo sapiens ACE (Stewart et al 1981;Skidgel et al 1984); …”

Section: Resultsmentioning

confidence: 99%

“…The enzyme is critical in the control of blood pressure and widely distributes in mammalian cells (Bakhle and Reynard 1971;Lee et al 1971a, b, c;Sander and Huggins 1971). However, at this moment, there have been only a few reports of microbial DCPs (Lee et al 1971d;Yaron et al 1972;Yaron 1976;Vimr and Miller 1983;Nagamori et al 1991;Miyoshi et al 1992;Ogasawara et al 1997;Rivière et al 2007). DCP is the only cytoplasmic C-terminal exopeptidase known for Escherichia coli (Yaron et al 1972;Yaron 1976).…”

Section: Introductionmentioning

confidence: 99%

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The dipeptidyl carboxypeptidase of Escherichia coli novablue: overproduction and molecular characterization of the recombinant enzyme

Chen

Chang

Lin

et al. 2008

World J Microbiol Biotechnol

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To express Escherichia coli novablue dipeptidyl carboxypeptidase (EcDCP), the gene was amplified by PCR and cloned into the expression plasmid pQE-31 to yield pQE-EcDCP. His 6 -tagged EcDCP (His 6 -EcDCP) was over-expressed in E. coli M15 (pQE-EcDCP) as a soluble and active form under 0.05 mM IPTG induction at 26°C for 12 h. The recombinant enzyme was purified to homogeneity by Ni 2? -NTA resin and had a molecular mass of approximately 75 kDa. The temperature and pH optima for His 6 -EcDCP were 37°C and 7.0, respectively. In the presence of 200 mM NaCl, His 6 -EcDCP was stimulated by 1.5 fold. The K M and k cat values of the enzyme for N-benzoyl-L-glycyl-L-histidyl-L-leucine were 1.83 mM and 168.3 s -1 , respectively. His 6 -EcDCP activity was dramatically inhibited by 10 mM EDTA, 0.25 mM 1.10-phenanthroline, and 2.5 mM DEPC, but it was not affected by Ser, Asp, Lys, and Trp protease inhibitors. Analysis of His 6 -EcDCP by circular dichroism revealed that the secondary structures of the enzyme in 30 mM universal buffer (pH 7.0) were 17% a-helix, 35% b-sheet and 47% random coil. Mid point of thermal transition was calculated to be 55°C for the recombinant enzyme.

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Section: Effects Of Metal Ions and Other Compounds On His 6 -Ecdcp Acsupporting

confidence: 91%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The dipeptidyl carboxypeptidase of Escherichia coli novablue: overproduction and molecular characterization of the recombinant enzyme

Chen

Chang

Lin

et al. 2008

World J Microbiol Biotechnol

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“…Apart from the functional homology between captopril and β-lactams, ACE is an evolutionary highly conserved protein. DNA sequence analysis revealed that ACE-homologos could be identified in a variety of different eukaryotic phyla and in procaryotes [220][221][222][223][224][225][226][227]. Some of these ACE-like enzymes have been purified and biochemically characterized in vitro.…”

Section: Inhibitors Of Angiotensin-converting Enzyme and Other Zinc-cmentioning

confidence: 99%

Are antibacterial effects of non-antibiotic drugs random or purposeful because of a common evolutionary origin of bacterial and mammalian targets?

Dalhoff

2020

Infection

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Purpose Advances in structural biology, genetics, bioinformatics, etc. resulted in the availability of an enormous pool of information enabling the analysis of the ancestry of pro- and eukaryotic genes and proteins. Methods This review summarizes findings of structural and/or functional homologies of pro- and eukaryotic enzymes catalysing analogous biological reactions because of their highly conserved active centres so that non-antibiotics interacted with bacterial targets. Results Protease inhibitors such as staurosporine or camostat inhibited bacterial serine/threonine or serine/tyrosine protein kinases, serine/threonine phosphatases, and serine/threonine kinases, to which penicillin-binding-proteins are linked, so that these drugs synergized with β-lactams, reverted aminoglycoside-resistance and attenuated bacterial virulence. Calcium antagonists such as nitrendipine or verapamil blocked not only prokaryotic ion channels but interacted with negatively charged bacterial cell membranes thus disrupting membrane energetics and inducing membrane stress response resulting in inhibition of P-glycoprotein such as bacterial pumps thus improving anti-mycobacterial activities of rifampicin, tetracycline, fluoroquinolones, bedaquilin and imipenem-activity against Acinetobacter spp. Ciclosporine and tacrolimus attenuated bacterial virulence. ACE-inhibitors like captopril interacted with metallo-β-lactamases thus reverting carbapenem-resistance; prokaryotic carbonic anhydrases were inhibited as well resulting in growth impairment. In general, non-antibiotics exerted weak antibacterial activities on their own but synergized with antibiotics, and/or reverted resistance and/or attenuated virulence. Conclusions Data summarized in this review support the theory that prokaryotic proteins represent targets for non-antibiotics because of a common evolutionary origin of bacterial- and mammalian targets resulting in highly conserved active centres of both, pro- and eukaryotic proteins with which the non-antibiotics interact and exert antibacterial actions.

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“…While zinc is an essential ion in many enzymes, e.g. proteases (Ogasawara et al 1997), cadmium is not essential for any organism. Cadmlum has been reported to cause a series of toxic effects, such as strong production of reactive oxygen species (Hassoun & Stohs 1996), depletion of glutathione (Shimizu et al 1997), inhibition of enzymes involved in DNA synthesis and repair (Yang et al 1996), and DNA single-strand breaks (Hassoun & Stohs 1996).…”

Section: Introductionmentioning

confidence: 99%

Accumulation of cadmium and zinc in the marine sponge Suberites domuncula and its potential consequences on single-strand breaks and on expression of heat-shock protein:a natural field study

Müller¹,

Batel²,

Lacorn³

et al. 1998

Mar. Ecol. Prog. Ser.

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The marine sponge Suberites domuncula (Porifera; Demospongiae) was used in a natural field study (in the vicinity of Rovinj, Croatia) as a bioindicator to assess the genotoxic risk of cadmium exposure for sponges as a model for invertebrates, and as an early stress marker of expression of heatshock protein 70 (M, 73 kDa; termed HSP73). Slgn~ficant differences in 'natural' cadmium levels of S. domunculd were found among sponges collected from differently polluted sites; in contrast zlnc levels were relatively similar. Sponges llving at polluted sites contained 15 to 24 times more cadmlum than sponges at unpolluted reference sites. The capacity of S. domuncula to accumulate cadmlun~ after an add~tlonal exposure to 5 mg I-' of cadmium for 5 d, was markedly higher (increase in cadmium content of 4 2 to 49 mg Cd kg-') for sponges collected from polluted sites compared to those from uncontaminated or less contaminated sites (increase of 8 to 29 mg Cd kg-'); a strong increase in cadmlum level was accompanied by a reduction in zinc level in the sponges from polluted sites. In this study we have used the rapid, sensitive and cheap Fast ~i c r o m e t h o d~ to determine the number of DNA single-stl-and breaks in S. domuncula collected from differently polluted sites and some of those sponges additionally exposed to cadmium. The strand scission factor (SSF) was found to depend on the pollution load; it further increased by 1.7-to 8.6-fold after additional exposure to cadmium (5 mg I-'). Similarly, the levels of HSP73 were found to be highest in S. domuncula living at polluted sites. Additional exposure of the sponges to cadmium strongly increased HSP73 level. This increase in HSP73 level was higher in sponges from polluted sites compared to those collected from uncontaminated or less contaminated control sites. From these data we conclude that the sponge S. domuncula is a useful bioindicator regarding the cadmium load of the aquatic environment.

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Purification and Characterization of a Dipeptidyl Carboxypeptidase from Pseudomonas sp. WO24

Cited by 9 publications

References 11 publications

The dipeptidyl carboxypeptidase of Escherichia coli novablue: overproduction and molecular characterization of the recombinant enzyme

The dipeptidyl carboxypeptidase of Escherichia coli novablue: overproduction and molecular characterization of the recombinant enzyme

Are antibacterial effects of non-antibiotic drugs random or purposeful because of a common evolutionary origin of bacterial and mammalian targets?

Accumulation of cadmium and zinc in the marine sponge Suberites domuncula and its potential consequences on single-strand breaks and on expression of heat-shock protein:a natural field study

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