Projects / Programmes
Analysis of interactions between inhibitors (stefins) and papain-like cysteine proteases
| Code |
Science |
Field |
Subfield |
| 1.05.00 |
Natural sciences and mathematics |
Biochemistry and molecular biology |
|
| Code |
Science |
Field |
| P004 |
Natural sciences and mathematics |
Biochemistry, Metabolism |
stefin, papain-like cysteine proteases, inhibition, 3-D structure, bimolecular interactions, AFM
Researchers (2)
| no. |
Code |
Name and surname |
Research area |
Role |
Period |
No. of publicationsNo. of publications |
| 1. |
19332 |
PhD Manca Kenig |
Biochemistry and molecular biology |
Researcher |
2003 - 2004 |
0 |
| 2. |
04988 |
PhD Dušan Turk |
Biochemistry and molecular biology |
Head |
2003 - 2004 |
0 |
Organisations (1)
| no. |
Code |
Research organisation |
City |
Registration number |
No. of publicationsNo. of publications |
| 1. |
0106 |
Jožef Stefan Institute |
Ljubljana |
5051606000 |
18 |
Abstract
The current understanding of mechanism of interaction between a cystatin-type inhibitor and a lysosomal cysteine protease is based on the crystal structures of chicken egg-white cystatin and the complex of human stefin B bound to carboxymethylated plant enzyme papain and kinetics data. We will attempt to systematically fill the gaps in the literature data and investigate the role of N-terminal residues in stefins. We will also address the question of long range interaction and the potential role of a protein fold: Is the fold of a stefin relevant only to present a compatible surface to the target protease or does it contribute to the binding also by long range interactions.
These will be achieved by combined use of protein engineering, kinetic measurements, X-ray crystallography and atom force microscopy. The enzyme kinetics studies (inhibition constant determination dependent on media composition including the various ionic strength of the media) will provide the basic assessment of the long range electrostatic interactions. With atom force microscopy acting forces between protease and inhibitor molecules will be measured. These measurements will enable us to estimate the magnitudes of the contact surface and the long range interactions in a bi-macromolecular reaction pathway. The knowledge of 3-dimensional structures of the complexes obtained by X-ray crystallography will be the key to design engineered proteins and interpret the kinetic and atom force microscopy data.
