ABSTRACT
The crystal structure of the H. influenzae YfeU protein, was determined at 1.90 A resolution using multi-wavelength anomalous diffraction. YfeU belongs to a very large conserved family of proteins found mainly in bacteria but also in archaea and eukaryota. The protein is a homolog of eukaryotic glucokinase regulator and is predicted to be a sugar phosphate isomerase or aminotransferase. Here we describe the structure of YfeU and discuss the possible function as an etherase possibly involved in peptidoglycan recycling.
Subject(s)
Bacterial Proteins/chemistry , Haemophilus influenzae/enzymology , Peptidoglycan/metabolism , Amino Acid Sequence , Binding Sites , Crystallography, X-Ray , Haemophilus influenzae/metabolism , Models, Molecular , Molecular Sequence Data , Protein Conformation , Protein Folding , Sequence AlignmentABSTRACT
The plant hormone indoleacetic acid (IAA or auxin) transcriptionally activates a select set of early genes. The Aux/IAA class of early auxin-responsive genes encodes a large family of short-lived, nuclear proteins. Aux/IAA polypeptides homo- and heterodimerize, and interact with auxin-response transcription factors (ARFs) via C-terminal regions conserved in both protein families. This shared region contains a predicted betaalphaalpha motif similar to the prokaryotic beta-ribbon DNA binding domain, which mediates both protein dimerization and DNA recognition. Here, we show by circular dichroism spectroscopy and by chemical cross-linking experiments that recombinant peptides corresponding to the predicted betaalphaalpha region of three Aux/IAA proteins from Arabidopsis thaliana contain substantial alpha-helical secondary structure and undergo homo- and heterotypic interactions in vitro. Our results indicate a similar biochemical function of the plant betaalphaalpha domain and suggest that the betaalphaalpha fold plays an important role in mediating combinatorial interactions of Aux/IAA and ARF proteins to specifically regulate secondary gene expression in response to auxin.
Subject(s)
Indoleacetic Acids/chemistry , Indoleacetic Acids/metabolism , Protein Folding , Amino Acid Sequence , Arabidopsis , Circular Dichroism , Molecular Sequence Data , Peptides/chemistry , Peptides/metabolism , Protein Structure, Secondary , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence AlignmentABSTRACT
Many small bacterial, archaebacterial, and eukaryotic genomes have been sequenced, and the larger eukaryotic genomes are predicted to be completely sequenced within the next decade. In all genomes sequenced to date, a large portion of these organisms' predicted protein coding regions encode polypeptides of unknown biochemical, biophysical, and/or cellular functions. Three-dimensional structures of these proteins may suggest biochemical or biophysical functions. Here we report the crystal structure of one such protein, MJ0577, from a hyperthermophile, Methanococcus jannaschii, at 1.7-A resolution. The structure contains a bound ATP, suggesting MJ0577 is an ATPase or an ATP-mediated molecular switch, which we confirm by biochemical experiments. Furthermore, the structure reveals different ATP binding motifs that are shared among many homologous hypothetical proteins in this family. This result indicates that structure-based assignment of molecular function is a viable approach for the large-scale biochemical assignment of proteins and for discovering new motifs, a basic premise of structural genomics.
Subject(s)
Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Carrier Proteins/chemistry , Carrier Proteins/genetics , Genome , Protein Structure, Secondary , Adenosine Triphosphatases/chemistry , Adenosine Triphosphatases/genetics , Adenosine Triphosphatases/metabolism , Adenosine Triphosphate/metabolism , Amino Acid Sequence , Bacterial Proteins/metabolism , Binding Sites , Carrier Proteins/metabolism , Computer Simulation , Conserved Sequence , Crystallography, X-Ray , Hydrogen Bonding , Methanococcus/genetics , Models, Molecular , Molecular Sequence Data , Sequence AlignmentABSTRACT
Deepwater rice can grow in the regions of Southeast Asia that are flooded during the monsoon season because it has several adaptations allowing it to survive under flooded conditions. One such adaptation is the ability for rapid internode elongation upon partial submergence to maintain its foliage above the rising flood water levels. Ethylene is considered to be the trigger of this growth response because deepwater conditions not only trap ethylene in submerged organs, but also enhance the activity of 1-aminocyclopropane-1-carboxylate (ACC) synthase. Herein we have studied the expression characteristics of two members of the five-member multigene family encoding ACC synthase in rice OS-ACS1 and OS-ACS2 and show that partial submergence induces expression of OS-ACS1 and suppresses expression of OS-ACS2. The induction of OS-ACS1 occurs within 12 h of partial submergence and at low oxygen concentrations. The data also suggest that deepwater conditions posttranscriptionally regulate ACC synthase activity. OS-ACS1 gene expression may contribute to longer-term ethylene production, but not to the initial, growth-promoting increase in ethylene synthesis.
Subject(s)
Genes, Plant , Isoenzymes/biosynthesis , Lyases/biosynthesis , Multigene Family , Oryza/physiology , DNA Primers , Enzyme Induction , Lyases/genetics , Oryza/enzymology , Oryza/genetics , Polymerase Chain Reaction , Seasons , Suppression, Genetic , WeatherSubject(s)
Ethylenes/metabolism , Genes, Plant/genetics , Plant Growth Regulators/metabolism , Plant Physiological Phenomena , Plant Proteins , Signal Transduction/genetics , Amino Acid Sequence , Cloning, Molecular , Conserved Sequence , Ethylenes/biosynthesis , Fruit/physiology , Gene Expression Regulation, Plant , Molecular Sequence Data , Plant Diseases , Plant Growth Regulators/biosynthesis , Plants/enzymology , Plants/genetics , Plants/metabolism , Receptors, Cell Surface/metabolismABSTRACT
The plant hormone ethylene is believed to be responsible for the ability of rice to grow in the deepwater regions of Southeast Asia. Ethylene production is induced by hypoxia, which is caused by flooding, because of enhanced activity of 1-aminocyclopropane-1-carboxylic acid (ACC) synthase, the key enzyme in the ethylene biosynthetic pathway. We have cloned three divergent members, (OS-ACS1, OS-ACS2, and OS-ACS3), of a multigene family encoding ACC synthase in rice. OS-ACS1 resides on chromosome 3 and OS-ACS3 on chromosome 5 in the rice genome. The OS-ACS1 and OS-ACS3 genes are induced by anaerobiosis and indoleacetic acid (IAA) + benzyladenine (BA) + LiCl treatment. The anaerobic induction is differential and tissue specific; OS-ACS1 is induced in the shoots, whereas OS-ACS3 is induced in the roots. These inductions are insensitive to protein synthesis inhibitors, suggesting that they are primary responses to the inducers. All three genes are actually induced when protein synthesis is inhibited, indicating that they may be under negative control or that their mRNAs are unstable. The OS-ACS1 gene was structurally characterized, and the function of its encoded protein (M(r) = 53 112 Da, pI 8.2) was confirmed by expression experiments in Escherichia coli. The protein contains all eleven invariant amino acid residues that are conserved between aminotransferases and ACC synthases cloned from various dicotyledonous plants. The amino acid sequence shares significant identity to other ACC synthases (69-34%) and is more similar to sequences in other plant species (69% with the tomato LE-ACS3) than to other rice ACC synthases (50-44%). The data suggest that the extraordinary degree of divergence among ACC synthase isoenzymes within each species arose early in plant evolution and before the divergence of monocotyledonous and dicotyledonous plants.
