The insertion of two amino acids into a transcriptional inducer converts it into a galactokinase

Platt, Adam; Ross, Helen C.; Hankin, Steven; Reece, Richard J.

doi:10.1073/pnas.97.7.3154

Cited by 109 publications

(112 citation statements)

References 25 publications

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“…Combined, these data suggest that although Gal1p is primarily a galactokinase, it can also function as a weak transcriptional inducer. This conclusion has also been supported through a number of in vitro experiments (9,13).…”

supporting

confidence: 50%

“…Gal3p is highly related to the galactokinases of the Leloir pathway, but the protein does not possess a galactokinase activity (8,9). It can, however, be converted into a galactokinase through the addition of two amino acids into its sequence (a serine and an alanine after Ser 164), (9) underscoring the similarity between it and the Leloir pathway enzyme.…”

mentioning

confidence: 99%

“…It can, however, be converted into a galactokinase through the addition of two amino acids into its sequence (a serine and an alanine after Ser 164), (9) underscoring the similarity between it and the Leloir pathway enzyme. The yeast galactokinase Gal1p shows 70% amino acid identity (Ͼ90% similarity) with Gal3p over its entire length.…”

mentioning

confidence: 99%

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Molecular Structure of Saccharomyces cerevisiae Gal1p, a Bifunctional Galactokinase and Transcriptional Inducer

Thoden

Sellick

Timson

et al. 2005

Journal of Biological Chemistry

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Gal1p of Saccharomyces cerevisiae is capable of performing two independent cellular functions. First, it is a key enzyme in the Leloir pathway for galactose metabolism where it catalyzes the conversion of ␣-D-galactose to galactose 1-phosphate. Second, it has the capacity to induce the transcription of the yeast GAL genes in response to the organism being challenged with galactose as the sole source of carbon. This latter function is normally performed by a highly related protein, Gal3p, but in its absence Gal1p can induce transcription, albeit inefficiently, both in vivo and in vitro. Here we report the x-ray structure of Gal1p in complex with ␣-D-galactose and Mg-adenosine 5-(␤,␥-imido)triphosphate (AMPPNP) determined to 2.4 Å resolution. Overall, the enzyme displays a marked bilobal appearance with the active site being wedged between distinct N-and C-terminal domains. Despite being considerably larger than other galactokinases, Gal1p shares a similar molecular architecture with these enzymes as well as with other members of the GHMP superfamily. The extraordinary levels of similarity between Gal1p and Gal3p (ϳ70% amino acid identity and ϳ90% similarity) have allowed a model for Gal3p to be constructed. By identifying the locations of mutations of Gal3p that result in altered transcriptional properties, we suggest potential models for Gal3p function and mechanisms for its interaction with the transcriptional inhibitor Gal80p. The GAL genetic switch has long been regarded as a paradigm for the control of gene expression in eukaryotes. Understanding the manner in which two of the proteins that function in transcriptional regulation interact with one another is an important step in determining the overall molecular mechanism of this switch.There are four enzymes in the Leloir pathway that are responsible for the conversion of ␤-D-galactose to the more metabolically useful glucose 1-phosphate (1). In the first step of this pathway, ␤-D-galactose is epimerized to ␣-D-galactose by galactose mutarotase. The following step involves the ATP-dependent addition of a phosphate group to the galactose C-1 hydroxyl by galactokinase to yield galactose 1-phosphate. Subsequently, a UMP moiety, derived from UDP-glucose, is transferred to galactose 1-phosphate to generate glucose 1-phosphate and UDPgalactose. This reaction is catalyzed by galactose-1-phosphate uridylyltransferase. To complete the pathway, UDP-galactose is converted to UDP-glucose by UDP-galactose 4-epimerase.

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Molecular Structure of Saccharomyces cerevisiae Gal1p, a Bifunctional Galactokinase and Transcriptional Inducer

Thoden

Sellick

Timson

et al. 2005

Journal of Biological Chemistry

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“…The relief of Gal80p's inhibition of Gal4p in the presence of galactose depends on the function of Gal3p. Gal3p is a paralog of Gal1p (4), the galactokinase and the first enzyme of the galactose utilization pathway, but Gal3p does not have galactokinase activity (5,6). Any one of several mutations dispersed in the Gal3p sequence (GAL3 constitutive mutant) causes the activation of GAL genes in the absence of galactose (7).…”

mentioning

confidence: 99%

Gene activation by interaction of an inhibitor with a cytoplasmic signaling protein

Peng

Hopper

2002

Proc. Natl. Acad. Sci. U.S.A.

105

127

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Galactose-inducible genes (GAL genes) in yeast Saccharomyces cerevisiae are efficiently transcribed only when the sequencespecific transcription activator Gal4p is activated. Activation of Gal4p requires the interaction between the Gal4p inhibitory protein Gal80p and the galactokinase paralog, Gal3p. It has been proposed that Gal3p binds to a Gal80p-Gal4p complex in the nucleus to activate Gal4p. Here, we present evidence that the Gal3p-Gal80p interaction occurs in the cytoplasm, and concurrently, Gal80p is removed from Gal4p at the GAL gene promoter. We also show that GAL gene expression can be activated by heterologous protein-protein interaction in the cytoplasm that is independent of galactose and Gal3p function. These results indicate that galactose-triggered Gal3p-Gal80p association in the cytoplasm activates Gal4p in the nucleus.

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“…However ScGal3 can be converted into a catalytically active galactokinase through the insertion of the two amino acids, Ser and Ala, into its sequence (4). Overexpression of ScGal1, or a phenomenon termed long-term adaptation, can recover a gal3 phenotype in S. cerevisiae (5).…”

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confidence: 99%

Induction of the gal Pathway and Cellulase Genes Involves No Transcriptional Inducer Function of the Galactokinase in Hypocrea jecorina

Hartl

Kubicek

Schink

2007

Journal of Biological Chemistry

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The Saccharomyces cerevisiae galactokinase ScGal1, a key enzyme for D-galactose metabolism, catalyzes the conversion of D-galactose to D-galactose 1-phosphate, whereas its catalytically inactive paralogue, ScGal3, activates the transcription of the GAL pathway genes. In Kluyveromyces lactis the transcriptional inducer function and the galactokinase activity are encoded by a single bifunctional KlGal1. Here, we investigated the cellular function of the single galactokinase GAL1 in the multicellular ascomycete Hypocrea jecorina ‫؍(‬ Trichoderma reesei) in the induction of the gal genes and of the galactokinase-dependent induction of the cellulase genes by lactose (1,4-O-␤-D-galactopyranosyl-D-glucose). A comparison of the transcriptional response of a strain deleted in the gal1 gene (no putative transcriptional inducer and no galactokinase activity), a strain expressing a catalytically inactive GAL1 version (no galactokinase activity but a putative inducer function), and a strain expressing the Escherichia coli galK (no putative transcriptional inducer but galactokinase activity) showed that, in contrast to the two yeasts, both the GAL1 protein and the galactokinase activity are fully dispensable for induction of the Leloir pathway gene gal7 by D-galactose and that only the galactokinase activity is required for cellulase induction by lactose. The data document a fundamental difference in the mechanisms by which yeasts and multicellular fungi respond to the presence of D-galactose, showing that the Gal1/Gal3-Gal4 -Gal80-dependent regulatory circuit does not operate in multicellular fungi.The enzymes of the Leloir pathway are responsible for the conversion of D-galactose to D-glucose 1-phosphate and have been identified in all biological kingdoms. In the model organism Saccharomyces cerevisiae, the genes encoding the three main Leloir pathway enzymes, GAL1 (galactose kinase), GAL7 (UDP-galactose uridylyltransferase), and GAL10 (UDP-galactose epimerase and aldose 1-epimerase), are clustered and coordinately regulated at the level of transcription. The S. cerevisiae GAL genes are repressed by D-glucose, expressed only at a very low basal level on other respiratory carbon sources, and highly induced (up to 1000-fold) when the cells are switched to a medium containing D-galactose. This transcriptional activation is mediated by the interplay of three proteins; in the presence of D-galactose and ATP, the transcriptional inducer ScGal3 associates with the ScGal80 repressor thereby alleviating its repressing effects. This allows the transcriptional activator ScGal4 to recruit the RNA polymerase II to each of the GAL genes (reviewed in Refs.1-3).The S. cerevisiae galactokinase ScGal1 displays a 73% amino acid identity to the ScGal3, but galactokinase activity is absent in ScGal3. However ScGal3 can be converted into a catalytically active galactokinase through the insertion of the two amino acids, Ser and Ala, into its sequence (4). Overexpression of ScGal1, or a phenomenon termed long-term adaptation, can recover a gal3 ph...

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The insertion of two amino acids into a transcriptional inducer converts it into a galactokinase

Cited by 109 publications

References 25 publications

Molecular Structure of Saccharomyces cerevisiae Gal1p, a Bifunctional Galactokinase and Transcriptional Inducer

Molecular Structure of Saccharomyces cerevisiae Gal1p, a Bifunctional Galactokinase and Transcriptional Inducer

Gene activation by interaction of an inhibitor with a cytoplasmic signaling protein

Induction of the gal Pathway and Cellulase Genes Involves No Transcriptional Inducer Function of the Galactokinase in Hypocrea jecorina

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