Amino acid and DNA sequences of an extracellular basic protease of <i>Dichelobacter nodosus</i> show that it is a member of the subtilisin family of proteases

Lilley, Glenn G.; Kortt, Alexander A.

doi:10.1111/j.1432-1033.1992.tb17385.x

Cited by 36 publications

(36 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, this enzyme has a longer carboxyl-terminal extension beyond the active site, which, together with the inserts in the catalytic region, makes the entire sequence almost 100 residues longer. With these structural properties, the StmPr1 protease is similar to the extracellular proteases of Xanthomonas campestris (19), D. nodosus (15), and Alteromonas sp. (20); the homology with these proteases is 49%, 40%, and 38% identity, respectively, for the mature proteins.…”

Section: Figmentioning

confidence: 83%

“…Amino-terminal sequencing of the 47-kDa band yielded the sequence LAPNDPYYQQ, which turned out to be absent from protein sequence data bases. The sequence, however, showed homology with the amino termini of several known bacterial proteases, the closest of which is a serine protease from Dichelobacter nodosus (15), a member of the family of subtilisin-like proteases (cf. Ref.…”

Section: Protease Purificationmentioning

confidence: 99%

See 1 more Smart Citation

The Major Extracellular Protease of the Nosocomial Pathogen Stenotrophomonas maltophilia

Windhorst

Frank

Georgieva

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

Stenotrophomonas maltophilia is increasingly emerging as a multiresistant pathogen in the hospital environment. In immunosuppressed patients, these bacteria may cause severe infections associated with tissue lesions such as pulmonary hemorrhage. This suggests proteolysis as a possible pathogenic mechanism in these infections. This study describes a protease with broad specificity secreted by S. maltophilia. The gene, termed StmPr1, codes for a 63-kDa precursor that is processed to the mature protein of 47 kDa. The enzyme is an alkaline serine protease that, by sequence homology and enzymic properties, can be further classified as a new member of the family of subtilases. It differs from the classic subtilisins in molecular size, in substrate specificity, and probably in the architecture of the active site. The StmPr1 protease is able to degrade several human proteins from serum and connective tissue. Furthermore, pan-protease inhibitors such as ␣ 1 -antitrypsin and ␣ 2 -macroglobulin were unable to abolish the activity of the bacterial protease. The data support the interpretation that the extracellular protease of S. maltophilia functions as a pathogenic factor and thus could serve as a target for the development of therapeutic agents.Stenotrophomonas maltophilia, formerly referred to as Xanthomonas maltophilia or Pseudomonas maltophilia (1, 2), is an aerobic nonfermentative Gram-negative bacterium of widespread occurrence. For healthy humans, it is regarded as an opportunistic germ; it has been implicated in a variety of infections without distinctive clinical features (for a review, see Ref.3). However, in immune-compromised patients, particularly those with bone marrow aplasia or receiving intensive chemotherapy, cases of fulminant hemorrhagic pneumonia have been reported, even with fatal outcome (4 -6). In patients not surviving infections with S. maltophilia, histological inspection of the lung tissue revealed massive bleeding caused by damage to the lung epithelium (4). There are further reports demonstrating involvement of this bacterium in massive hemorrhagic processes of the small intestine and the subclavian artery accompanied by severe lesions of the tissue (5, 6). These observations strongly suggest the participation of proteolytic activity, produced by the bacteria, which may damage the infected tissue. Indeed, it is known that members of the Pseudomonaceae express and secrete a variety of proteases (cf. Ref. 7). Whereas the primary function of these enzymes is to provide a source of free amino acids for bacterial survival and growth, there is accumulating evidence that bacterial proteases may play a pathogenic role in the infected host by involvement in tissue invasion and destruction, evasion of host defenses, and modulation of the host immune system (8).The broad administration of antibiotics currently applied in cases of intensive care patients leads to selection of multiresistant S. maltophilia strains. Consequently, these bacteria are found with increasing frequency in the hospital environmen...

show abstract

Section: Figmentioning

confidence: 83%

Section: Protease Purificationmentioning

confidence: 99%

The Major Extracellular Protease of the Nosocomial Pathogen Stenotrophomonas maltophilia

Windhorst

Frank

Georgieva

et al. 2002

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…BprV is one of the three extracellular serine proteases of D. nodosus and has a predicted pI of 9.5 (36). Since in other bacteria Fur has been shown to regulate the production of secreted proteins (16,60), the secreted proteins from D. nodosus were analyzed over a pH 3 to 10 (not pH 4 to 7) range so that any potential effects on BprV could be detected, in addition to the two other acidic proteases.…”

Section: Resultsmentioning

confidence: 99%

Identification of a Dichelobacter nodosus Ferric Uptake Regulator and Determination of Its Regulatory Targets

Parker

Kennan

Myers³

et al. 2005

J Bacteriol

View full text Add to dashboard Cite

The expression of iron regulated genes in bacteria is typically controlled by the ferric uptake regulator (Fur) protein, a global transcriptional repressor that regulates functions as diverse as iron acquisition, oxidative stress, and virulence. We have identified a fur homologue in Dichelobacter nodosus, the causative agent of ovine footrot, and shown that it complements an Escherichia coli fur mutant. Homology modeling of the D. nodosus Fur protein with the recently solved crystal structure of Fur from Pseudomonas aeruginosa indicated extensive structural conservation. As Southern hybridization analysis of different clinical isolates of D. nodosus indicated that the fur gene was present in all of these strains, the fur gene was insertionally inactivated to determine its functional role. Analysis of these mutants by various techniques did not indicate any significant differences in the expression of known virulence genes or in iron-dependent growth. However, we determined several Fur regulatory targets by two-dimensional gel electrophoresis coupled with mass spectrometry. Analysis of proteins from cytoplasmic, membrane, and extracellular fractions revealed numerous differentially expressed proteins. The transcriptional basis of these differences was analyzed by using quantitative reverse transcriptase PCR. Proteins with increased expression in the fur mutant were homologues of the periplasmic iron binding protein YfeA and a cobalt chelatase, CbiK. Down-regulated proteins included a putative manganese superoxide dismutase and ornithine decarboxylase. Based on these data, it is suggested that in D. nodosus the Fur protein functions as a regulator of iron and oxidative metabolism.

show abstract

“…Skim Plasmid construction. pBR3I<B, a derivative of pBR322 containing the 3 kb HindIII-EcoRI fragment of the bprV gene (Lilley et a/., 1992), was used as the starting plasmid in this study. pCP95 and pCP97 were derived from pBR3KB a; follows.…”

Section: Methodsmentioning

confidence: 99%

Expression in Escherichia coli of the extracellular basic protease from Dichelobacter nodosus

Vaughan

Wang²,

Lilley³

et al. 1994

Microbiology

View full text Add to dashboard Cite

Dichelobacter nodosus, a Gram-negative obligate anaerobe and the causative agent of ovine footrot, secretes a number of extracellular proteases, one of which is highly basic in nature. The gene (bprV) encoding this basic protease, from virulent strain 198, has been cloned and sequenced. Clone pBR3KB contained the complete bprV gene which constitutively expressed an active protease using its own promoter, when cloned in Escherichia coli. However, levels of protease expression were low and unstable when the clone was expressed in liquid culture. A range of E. coli strains were examined for stable expression; strains NH274 and SURE@ were found to be better hosts for stable expression than other commonly used E. coli host strains. Stabilization and enhancement of expression was achieved by deletion of the native promoter region and expression from plasmid promoter or promoters, and by modification of culture conditions. The recombinant protease obtained from E. coli was indistinguishable from the native enzyme in size, activity, isoelectric point and immunological properties.

show abstract

Amino acid and DNA sequences of an extracellular basic protease of Dichelobacter nodosus show that it is a member of the subtilisin family of proteases

Cited by 36 publications

References 44 publications

The Major Extracellular Protease of the Nosocomial Pathogen Stenotrophomonas maltophilia

The Major Extracellular Protease of the Nosocomial Pathogen Stenotrophomonas maltophilia

Identification of a Dichelobacter nodosus Ferric Uptake Regulator and Determination of Its Regulatory Targets

Expression in Escherichia coli of the extracellular basic protease from Dichelobacter nodosus

Contact Info

Product

Resources

About