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Trypanosoma brucei thymidine kinase is tandem protein consisting of two homologous parts, which together enable efficient substrate binding.

ABSTRACT: Trypanosoma brucei causes African sleeping sickness, a disease for which existing chemotherapies are limited by their toxicity or lack of efficacy. We have found that four parasites, including T. brucei, contain genes where two or four thymidine kinase (TK) sequences are fused into a single open reading frame. The T. brucei full-length enzyme as well as its two constituent parts, domain 1 and domain 2, were separately expressed and characterized. Of potential interest for nucleoside analog development, T. brucei TK was less discriminative against purines than human TK1 with the following order of catalytic efficiencies: thymidine > deoxyuridine ? deoxyinosine > deoxyguanosine. Proteins from the TK1 family are generally dimers or tetramers, and the quaternary structure is linked to substrate affinity. T. brucei TK was primarily monomeric but can be considered a two-domain pseudodimer. Independent kinetic analysis of the two domains showed that only domain 2 was active. It had a similar turnover number (k(cat)) as the full-length enzyme but could not self-dimerize efficiently and had a 5-fold reduced thymidine/deoxyuridine affinity. Domain 1, which lacks three conserved active site residues, can therefore be considered a covalently attached structural partner that enhances substrate binding to domain 2. A consequence of the non-catalytic role of domain 1 is that its active site residues are released from evolutionary pressure, which can be advantageous for developing new catalytic functions. In addition, nearly identical 89-bp sequences present in both domains suggest that the exchange of genetic material between them can further promote evolution.

SUBMITTER: Ranjbarian F

PROVIDER: S-EPMC3366807 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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Trypanosoma brucei thymidine kinase is tandem protein consisting of two homologous parts, which together enable efficient substrate binding.

Ranjbarian Farahnaz F Vodnala Munender M Vodnala Sharvani Munender SM Rofougaran Reza R Thelander Lars L Hofer Anders A

The Journal of biological chemistry 20120321 21

Trypanosoma brucei causes African sleeping sickness, a disease for which existing chemotherapies are limited by their toxicity or lack of efficacy. We have found that four parasites, including T. brucei, contain genes where two or four thymidine kinase (TK) sequences are fused into a single open reading frame. The T. brucei full-length enzyme as well as its two constituent parts, domain 1 and domain 2, were separately expressed and characterized. Of potential interest for nucleoside analog devel ...[more]

PMID: 22442154

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Project description:BackgroundThe genome sequence of the protistan parasite Trypanosoma brucei contains many tandem gene arrays. Gene duplicates are created through tandem duplication and are expressed through polycistronic transcription, suggesting that the primary purpose of long, tandem arrays is to increase gene dosage in an environment where individual gene promoters are absent. This report presents the first account of the tandem gene arrays in the T. brucei genome, employing several related genome sequences to establish how variation is created and removed.ResultsA systematic survey of tandem gene arrays showed that substantial sequence variation existed across the genome; variation from different regions of an array often produced inconsistent phylogenetic affinities. Phylogenetic relationships of gene duplicates were consistent with concerted evolution being a widespread homogenising force. However, tandem duplicates were not usually identical; therefore, any homogenising effect was coincident with divergence among duplicates. Allelic gene conversion was detected using various criteria and was apparently able to both remove and introduce sequence variation. Tandem arrays containing structural heterogeneity demonstrated how sequence homogenisation and differentiation can occur within a single locus.ConclusionThe use of multiple genome sequences in a comparative analysis of tandem gene arrays identified substantial sequence variation among gene duplicates. The distribution of sequence variation is determined by a dynamic balance of conservative and innovative evolutionary forces. Gene trees from various species showed that intraspecific duplicates evolve in concert, perhaps through frequent gene conversion, although this does not prevent sequence divergence, especially where structural heterogeneity physically separates a duplicate from its neighbours. In describing dynamics of sequence variation that have consequences beyond gene dosage, this survey provides a basis for uncovering the hidden functionality within tandem gene arrays in trypanosomatids.

Dataset Information

Trypanosoma brucei thymidine kinase is tandem protein consisting of two homologous parts, which together enable efficient substrate binding.

Publications

Trypanosoma brucei thymidine kinase is tandem protein consisting of two homologous parts, which together enable efficient substrate binding.

Similar Datasets

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