-
Izražanje in izolacija rekombinantnih človeških katepsinov O in F : magistrsko deloFonović, MarkoRecombinant human cathepsins O and F were obtained from geneticaly modified bacterial and yeast cells. Yeast P. pastories, with human cathepsin F DNA inserted in its genome (yeast strain was provided ... by dr. Dieter Bromme, Mount Sinai School of Medicine, New York) was grown in fermentor. Expression of cathepsin F was induced by addition of methanol in the growth medium. Recombinant cathepsin F was isolated from fermentation broth with hydrophobic and gel chromatography. Protein identity was confirmed by immunological detection, enzyme activity measurement and N-terminal aminoacid analysis. Protein purity and homogenity was checked by SDS-PAGE and PAGE under native conditions. Isolation of cathepsin F was very troublesome and it was suspectedthat this is due to overglycosilation of the protein. In order to avoid this problem a mutant of cathepsin F cDNA without N-glycosilation sites,was prepared. Desired mutations were confirmed by DNA sequence analysis.Mutant cDNA cloned in expression plasmids pPIC9 and pGAPZalfa and yeast P. pastoris was transformed with both plasmids. After test expression nomutant cathepsin F was detected in the growth medium. No mutant protein was expressed in yeast cells. Procathepsin O cDNA was inserted in expression plasmid pGAPZalfa and P.pastoris strain X-33 was transformed by constructed plasmid. Transformed yeast cells were grown in shaker flasks and protein expression was controled by SDS-PAGE. After 4-6 days of expression a protein band with molecular mass similar to procathepsin O, was observed. Presence of procathepsin O was confirmed with N-terminal aminoacid sequence analysis. Procathepsin O was partially purified by gel chromatography. Activation at acidic pH was performed and monitored by SDS-PAGE. (Abstract truncated at 2000 characters).Type of material - master's thesisPublication and manufacture - Ljubljana : [M. Fonovič], 2001Language - slovenianCOBISS.SI-ID - 2317844
Author
Fonović, Marko
Other authors
Turk, Vito |
Turk, Boris, 1964-
Topics
Cathepsins |
Isolation and purification |
Cathepsins |
Genetics |
Cathepsins |
Physiology |
Recombinant proteins |
Katepsini |
Izolacija in čiščenje |
Katepsini |
Genetika |
Katepsini |
Fiziologija |
Rekombinantne beljakovine |
biokemija |
molekularna biologija
![loading ... loading ...](themes/default/img/ajax-loading.gif)
Library/institution |
City | Acronym | For loan | Other holdings |
---|---|---|---|---|
MF, Central Medical Library, Ljubljana | Ljubljana | CMK |
outside loan 1 cop.
|
![loading ... loading ...](themes/default/img/ajax-loading.gif)
![loading ... loading ...](themes/default/img/ajax-loading.gif)
![loading ... loading ...](themes/default/img/ajax-loading.gif)
Shelf entry
Permalink
- URL:
Impact factor
Access to the JCR database is permitted only to users from Slovenia. Your current IP address is not on the list of IP addresses with access permission, and authentication with the relevant AAI accout is required.
Year | Impact factor | Edition | Category | Classification | ||||
---|---|---|---|---|---|---|---|---|
JCR | SNIP | JCR | SNIP | JCR | SNIP | JCR | SNIP |
Select the library membership card:
DRS, in which the journal is indexed
Database name | Field | Year |
---|
Links to authors' personal bibliographies | Links to information on researchers in the SICRIS system |
---|---|
Fonović, Marko | 18801 |
Turk, Vito | 01085 |
Turk, Boris, 1964- | 07561 |
Select pickup location:
Material pickup by post
Notification
Subject headings in COBISS General List of Subject Headings
Select pickup location
Pickup location | Material status | Reservation |
---|
Please wait a moment.