Protein Refinement and Engineering


Our research is focussed on the exciting interface between chemistry and biology. This area of research is crucial for understanding both the molecular basis of biological phenomena and for the utilisation of biological materials. Projects usually start with structure determination using diffraction techniques. Following this, structures are used to rationalise protein behaviour and to design experiments that further probe the intimate relationship between sequence, structure and function. Our current work has focussed on the following systems: 1) signal transduction: the GlnK and PII protein; 2) cell surface receptors: the b subunit of the Il5 receptor; and 3) replication protein: the e subunit of DNA polymerase III.


Principal Investigator

David Ollis
Research School of Chemistry
ANU

Project

s06

Facilities Used

SC, PC

Co-Investigator

Paul Carr
Research School of Chemistry
ANU

RFCD Codes

270108, 270199, 270805


Significant Achievements, Anticipated Outcomes and Future Work

The main experimental technique used by the group is X-ray diffraction. The object of our research is to firstly determine the structures of large bio-molecules and to use this information to understand how the molecules function.

The structure of the extra-cellular component of the b common receptor of the IL5 cytokine was published during 2001. IL5 plays a role in the generation of eosinophilia characteristic of helminth infections and allergic diseases such as asthma. The receptor is vital to understanding how IL5 stimulates cells. The arrangement of domains within the receptor is novel and allowed interaction sites with IL5 to be predicted. Experiments to verify these predictions are in progress.

The structure of the PII protein as a complex with ATP was also published during the year. Previous papers have described the protein structure while the latest manuscript describes the interaction between the protein and ATP, a co-factor that is necessary for biological activity.

 

Computational Techniques Used

Electron density maps or model construction are produced by Fourier techniques. Models are refined with non-linear least squares programs that minimise the difference between the observed and calculated structure amplitudes.

 

Publications, Awards and External Funding

Carr, P.D., Verger, D., Ashton, A.R. and Ollis, D.L. Chloroplast NADP-malate dehydrogenase: structural basis of light-dependent regulation of activity by thiol oxidation and reduction. Structure with folding and design (1999) 7 461- 475.

Lappanen, VL., Merckel, M., Ollis, D.L., Wong, K.K., Kozarach, J.W, and Goldman, A. Pyruvate formate lyase is structurally homologous, to type I ribonucleotide reductase. Structure with folding and design (1999) 7 733-744.

Ashton, A.R., Trevanion, S.J. , Carr, P.D., Verger D. and Ollis, D.L. Structural basis for the light regulation of chloroplast NADP malate dehydrogenase Physiologia. Plantarum, (2000) 110, 314-321.

Robinson, A., Edwards, K., Carr, P.D., Barton, J.D. Gary. D. Ewart and Ollis, D.L. Structure of the C123S mutant of dienelactone hydrolase (DLH) bound with the PMS moiety of the protease inhibitor phenylmethylsulfonyl fluoride (PMSF). Acta Cryst D (2000) D56 1376-1384.

Carr, P.D., Gustin, S.E., Church, A.P., Murphy, J.M., Ford, S.C., Mann, D.A., Woltring, D.M., Walker, I., Ollis, D.L. amd Young, I.G. Structure of the complete extracellular domain of the common b subunit of the human GM-CSF, IL-3 and IL-5 receptors reveals a novel dimer configuration. Cell (2001) 104, 291-300.

Xu, Y., Carr, P.D., Huber, T., Vasudevan, S.G. and Ollis, D.L. The structure of the PII - ATP complex. European Journal of Biochemistry (2001) 268, 2028 -2037.

Hamdan, S., Carr, P.D., Brown, S.E., Ollis, D.L., Dixon, N.E. Structural basis for proofreading during replication of the Escheria coli chromosome. Structure with folding and design (in press).