Colicin production in Escherichia coli (E. coli) strains represents an important trait with regard to microbial survival and competition in the complex intestinal environment. A novel colicin type, colicin Z (26.3 kDa), was described as a product of an original producer, extraintestinal E. coli B1356 strain, isolated from the anorectal abscess of a 17 years-old man. The 4,007 bp plasmid (pColZ) was completely sequenced and colicin Z activity (cza) and colicin Z immunity (czi) genes were identified. The cza and czi genes are transcribed in opposite directions and encode for 237 and 151 amino acid-long proteins, respectively. Colicin Z shows a narrow inhibitory spectrum, being active only against enteroinvasive E. coli (EIEC) and Shigella strains via CjrC receptor recognition and CjrB- and ExbB-, ExbD-mediated colicin translocation. All tested EIEC and Shigella strains isolated between the years 1958-2010 were sensitive to colicin Z. The lethal effect of colicin Z was found to be directed against cell wall peptidoglycan (PG) resulting in PG degradation, as revealed by experiments with Remazol Brilliant Blue-stained purified peptidoglycans and with MALDI-TOF MS analyses of treated PG. Colicin Z represents a new class of colicins that is structurally and functionally distinct from previously studied colicin types.

• MeSH
• Escherichia coli genetika   MeSH
• koliciny genetika   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• mladiství MeSH
• plazmidy genetika   MeSH
• sekvence nukleotidů MeSH
• Shigella genetika   MeSH
• Check Tag
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• koliciny MeSH

Turbidimetric method with spectrophotometric detection of changes in density of test bacteria S. aureus strain SA 812 for determination of bacteriolytic activity of lysostaphin was employed. Results of two evaluations are compared: (1) calculation of the relative value of turbidity decrease on the basis of the difference of absolute values of A540 at the beginning of reaction and after the incubation period, (2) following of time changes in A540 by monitoring the course of reaction directly in the constant-temperature cuvette of the spectrophotometer at 37 degrees C. Both arrangements yielded identical results, within the significance level of 0.05. With concentrated samples both methods yield reliable results; with diluted samples the accuracy of the "absolute" method decreases together with decreasing lysostaphin concentration much faster than with the "registration" method. The registration method makes it possible to detect even minute amounts of the lytic enzyme and thus to distinguish the values of activity in dilute samples even when data obtained by means of the "absolute" method cannot be considered as reliable. A unit of bacteriolytic activity can be expressed from the kinetic curve as an amount of enzyme preparation causing delta A540/min = 0.01.

• MeSH
• bakteriolýza účinky léků   MeSH
• fluorometrie MeSH
• lysostafin farmakologie   MeSH
• nefelometrie a turbidimetrie * metody   MeSH
• spektrofotometrie * metody   MeSH
• Staphylococcus aureus účinky léků   MeSH
• teplota MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• lysostafin MeSH

The present paper reports a modified method for isolation of lysostaphin--a bacteriolytic agent with specific affinity for staphylococcal cell wall. The proposed purification scheme includes three steps. The first procedure is ultrafiltration through a membrane filter giving a yield of 75.6%. The result of ultrafiltration is a concentrated, 10-times purified preparation of lysostaphin with specific activity 0.62 U/mg which can be used for digestion of S. aureus cells. Further step, performed by ion-exchange chromatography on DEAE-cellulose, yields a 60-times purified preparation containing a mixture of enzyme components of lysostaphin. The yield of this step is 47.2%, the preparation contains 3.54 U/mg protein. Using gel filtration on Sephadex G-50 a component with hexosaminidase activity was separated from the endopeptidase component on the basis of molar mass difference. A 270-times purified preparation of lysostaphin-endopeptidase with minimum of contaminating substances was obtained in this step. The yield of gel filtration was 22.1%, specific activity increased up to 16.3 U/mg protein.

• MeSH
• chromatografie iontoměničová MeSH
• filtrace MeSH
• gelová chromatografie MeSH
• kultivační média MeSH
• lysostafin izolace a purifikace   MeSH
• mikrobiologické techniky MeSH
• molekulární sekvence - údaje MeSH
• sacharidové sekvence MeSH
• sekvence aminokyselin MeSH
• Staphylococcus chemie   růst a vývoj   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kultivační média MeSH
• lysostafin MeSH