Dataset Information

Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline.

ABSTRACT: Structural genomics is emerging as a principal approach to define protein structure-function relationships. To apply this approach on a genomic scale, novel methods and technologies must be developed to determine large numbers of structures. We describe the design and implementation of a high-throughput structural genomics pipeline and its application to the proteome of the thermophilic bacterium Thermotoga maritima. By using this pipeline, we successfully cloned and attempted expression of 1,376 of the predicted 1,877 genes (73%) and have identified crystallization conditions for 432 proteins, comprising 23% of the T. maritima proteome. Representative structures from TM0423 glycerol dehydrogenase and TM0449 thymidylate synthase-complementing protein are presented as examples of final outputs from the pipeline.

SUBMITTER: Lesley SA

PROVIDER: S-EPMC129326 | biostudies-literature | 2002 Sep

REPOSITORIES: biostudies-literature

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Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline.

Lesley Scott A SA Kuhn Peter P Godzik Adam A Deacon Ashley M AM Mathews Irimpan I Kreusch Andreas A Spraggon Glen G Klock Heath E HE McMullan Daniel D Shin Tanya T Vincent Juli J Robb Alyssa A Brinen Linda S LS Miller Mitchell D MD McPhillips Timothy M TM Miller Mark A MA Scheibe Daniel D Canaves Jaume M JM Guda Chittibabu C Jaroszewski Lukasz L Selby Thomas L TL Elsliger Marc-Andre MA Wooley John J Taylor Susan S SS Hodgson Keith O KO Wilson Ian A IA Schultz Peter G PG Stevens Raymond C RC

Proceedings of the National Academy of Sciences of the United States of America 20020822 18

Structural genomics is emerging as a principal approach to define protein structure-function relationships. To apply this approach on a genomic scale, novel methods and technologies must be developed to determine large numbers of structures. We describe the design and implementation of a high-throughput structural genomics pipeline and its application to the proteome of the thermophilic bacterium Thermotoga maritima. By using this pipeline, we successfully cloned and attempted expression of 1,37 ...[more]

PMID: 12193646

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Project description:Structural genomic projects envision almost routine protein structure determinations, which are currently imaginable only for small proteins with molecular weights below 25,000 Da. For larger proteins, structural insight can be obtained by breaking them into small segments of amino acid sequences that can fold into native structures, even when isolated from the rest of the protein. Such segments are autonomously folding units (AFU) and have sizes suitable for fast structural analyses. Here, we propose to expand an intuitive procedure often employed for identifying biologically important domains to an automatic method for detecting putative folded protein fragments. The procedure is based on the recognition that large proteins can be regarded as a combination of independent domains conserved among diverse organisms. We thus have developed a program that reorganizes the output of BLAST searches and detects regions with a large number of similar sequences. To automate the detection process, it is reduced to a simple geometrical problem of recognizing rectangular shaped elevations in a graph that plots the number of similar sequences at each residue of a query sequence. We used our program to quantitatively corroborate the premise that segments with conserved sequences correspond to domains that fold into native structures. We applied our program to a test data set composed of 99 amino acid sequences containing 150 segments with structures listed in the Protein Data Bank, and thus known to fold into native structures. Overall, the fragments identified by our program have an almost 50% probability of forming a native structure, and comparable results are observed with sequences containing domain linkers classified in SCOP. Furthermore, we verified that our program identifies AFU in libraries from various organisms, and we found a significant number of AFU candidates for structural analysis, covering an estimated 5 to 20% of the genomic databases. Altogether, these results argue that methods based on sequence similarity can be useful for dissecting large proteins into small autonomously folding domains, and such methods may provide an efficient support to structural genomics projects.

Dataset Information

Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline.

Publications

Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline.

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