Partner und Internationale Organisationen
(Englisch)
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Karolinska Institutet Stockholm (S), Universitat de Barcelona (E), Universitat Autonoma de Barcelona (E), Philipps Universität Marburg (D)
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Abstract
(Englisch)
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The growing number of characterised sequences from the human genome project, other genome studies, and analyses of individual genes, cDNAs and gene products, offer great potentials to understand:
· gene products, how they are folded and functionally active, i.e. promoting the era of proteome research.
· molecular evolution, with duplications, mutations and rearrange-ments,
· biotechnology and its possibilities to correct diseases, and to promote industrial pro-duction
In order to fulfill these objectives, our programme aims at:
· Sequence data searches with algorithms for alignments, homology assessments, corre-lations with gene borders, protein structures, and conformational folds.
· Sequence analysis of novel cDNAs or proteins, and corresponding functional analysis.
· Use of these combined 'in silico' and 'wet chemistry' data to analyse the gene families thus obtained to understand functions of the corresponding products, and to include muta-genesis and expressions as well as species variants to check all conclu-sions.
Short-term goals of the project involve to screen all novel genomes emerging, existing data-banks, and novel gene products defined at the protein, cDNA, or genome level. Emphasis will be on dehydrogenases, but with consi-derable efforts also on peptide hormones, membrane proteins and proteases. The work will center on definitions of general interest and function. Comparisons of known structures will define motifs, which will be used for further scree-nings, complemented with mutagenesis and molecular modelling to check conclusions and to functionally characterise novel proteins within the families. The increased knowledge will give products (clones, gene pro-ducts, cell lines, computer programmes, databases) and pa-tents, and will allow construction of new products for medical applications and pharmaceu-tical use.
Work in our group was centered around the study of structure-function rela-tionships and evolutionary aspects of carbonyl reductases and related proteins. Using the human enzyme as a model we identified a neutral and two basic residues in the N-terminal part which are important for coenzyme binding and a cysteine residue in the C-terminal part required for substrate binding. In addition, we determined the structural requirements permitting the autocatalytic modification by oxocarboxylic acids of a lysine residue at the entrance of the substrate binding cleft and showed that substitution of glutamate for the homo-logous lysine is responsible for differences in substrate specificity between two closely related rat carbonyl reductases.
By combining 'in silico' studies with 'wet chemistry' we demonstrated that carbonyl reductase from Branchio-stoma floridae exhibits many characteristics of the mammalian enzymes, whereas a carbonyl reductase-like protein from Arabidopsis thaliana did not cross-react with antibodies against human carbonyl reductase and was essentially inactive using standard substrates of mammalian carbonyl reductases.
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