A low-background-intensity focusing small-angle X-ray scattering undulator beamline

Kirby, Nigel; Mudie, Stephen; Hawley, Adrian; Cookson, David J.; Mertens, Haydyn D. T.; Cowieson, Nathan; Samardzic-Boban, Vesna

doi:10.1107/s002188981302774x

Cited by 482 publications

(452 citation statements)

References 16 publications

Supporting

Mentioning

449

Contrasting

Order By: Relevance

“…SAXS data were collected at the Australian Synchrotron SAXS/WAXS beamline [23]. The wavelength of the X-rays was 1.0332 A and the sample detector distance was 2600 mm.…”

Section: Saxs Data Collectionmentioning

confidence: 99%

Independent catalysis of the short form HisG from Lactococcus lactis

et al. 2016

View full text Add to dashboard Cite

Edited by Michael IbbaATP-phosphoribosyltransferase (ATP-PRT) catalyses the first step of histidine biosynthesis. Two different forms of ATP-PRT have been described; the homo-hexameric long form, and the hetero-octameric short form. Lactococcus lactis possesses the short form ATP-PRT comprising four subunits of HisG S , the catalytic subunit, and four subunits of HisZ, a histidyl-tRNA synthetase paralogue. Previous studies have suggested that HisG S requires HisZ for catalysis. Here, we reveal that the dimeric HisG S does display ATP-PRT activity in the absence of HisZ. This result reflects the evolutionary relationship between the long and short form ATP-PRT, which acquired allosteric inhibition and enhanced catalysis via two divergent strategies.

show abstract

“…SAXS data were collected at the Australian Synchrotron SAXS/WAXS beamline [23]. The wavelength of the X-rays was 1.0332 A and the sample detector distance was 2600 mm.…”

Section: Saxs Data Collectionmentioning

confidence: 99%

Independent catalysis of the short form HisG from Lactococcus lactis

et al. 2016

View full text Add to dashboard Cite

show abstract

“…To examine the relationship between the full-length crystal structure and the oligomers of PAH that exist in solution in the absence of Phe, we performed SAXS analysis on the PAH preparations used for crystallography using size-exclusion chromatography in-line with synchrotron SAXS (20). The experimental radius of gyration (R g ), maximum interatomic distance (D max ) (Tables S3 and S4), and pairwise shape distribution (Fig.…”

Section: Saxs Correlates the Pah Crystal Structure With The Solution mentioning

confidence: 99%

First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer

Arturo

Gupta

Héroux

et al. 2016

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

122

View full text Add to dashboard Cite

Improved understanding of the relationship among structure, dynamics, and function for the enzyme phenylalanine hydroxylase (PAH) can lead to needed new therapies for phenylketonuria, the most common inborn error of amino acid metabolism. PAH is a multidomain homo-multimeric protein whose conformation and multimerization properties respond to allosteric activation by the substrate phenylalanine (Phe); the allosteric regulation is necessary to maintain Phe below neurotoxic levels. A recently introduced model for allosteric regulation of PAH involves major domain motions and architecturally distinct PAH tetramers [Jaffe EK, Stith L, Lawrence SH, Andrake M, Dunbrack RL, Jr (2013) Arch Biochem Biophys 530(2):73-82]. Herein, we present, to our knowledge, the first X-ray crystal structure for a full-length mammalian (rat) PAH in an autoinhibited conformation. Chromatographic isolation of a monodisperse tetrameric PAH, in the absence of Phe, facilitated determination of the 2.9 Å crystal structure. The structure of full-length PAH supersedes a composite homology model that had been used extensively to rationalize phenylketonuria genotype-phenotype relationships. Small-angle X-ray scattering (SAXS) confirms that this tetramer, which dominates in the absence of Phe, is different from a Phestabilized allosterically activated PAH tetramer. The lack of structural detail for activated PAH remains a barrier to complete understanding of phenylketonuria genotype-phenotype relationships. Nevertheless, the use of SAXS and X-ray crystallography together to inspect PAH structure provides, to our knowledge, the first complete view of the enzyme in a tetrameric form that was not possible with prior partial crystal structures, and facilitates interpretation of a wealth of biochemical and structural data that was hitherto impossible to evaluate.phenylalanine hydroxylase | phenylketonuria | X-ray crystallography | small-angle X-ray scattering | allosteric regulation M ammalian phenylalanine hydroxylase (PAH) (EC 1.14.16.1) is a multidomain homo-multimeric protein whose dysfunction causes the most common inborn error in amino acid metabolism, phenylketonuria (PKU), and milder forms of hyperphenylalaninemia (OMIM 261600) (1). PAH catalyzes the hydroxylation of phenylalanine (Phe) to tyrosine, using nonheme iron and the cosubstrates tetrahydrobiopterin and molecular oxygen (2, 3). A detailed kinetic mechanism has recently been derived from elegant single-turnover studies (4). PAH activity must be carefully regulated, because although Phe is an essential amino acid, high Phe levels are neurotoxic. Thus, Phe allosterically activates PAH by binding to a regulatory domain. Phosphorylation at Ser16 potentiates the effects of Phe, with phosphorylated PAH achieving full activation at lower Phe concentrations than the unphosphorylated protein (5, 6). Allosteric activation by Phe is accompanied by a major conformational change, as evidenced by changes in protein fluorescence and proteolytic susceptibility, and by stabilization of a tetrameric confo...

show abstract

“…The detailed beamline properties are described in Kirby et al (2013). Samples were prepared as described above, placed into 1.5-mm-diameter quartz capillaries, and sealed with epoxy resin.…”

Section: Saxs and Waxs Experimentsmentioning

confidence: 99%

Disordered Cold Regulated15 Proteins Protect Chloroplast Membranes during Freezing through Binding and Folding, But Do Not Stabilize Chloroplast Enzymes in Vivo

et al. 2014

View full text Add to dashboard Cite

Freezing can severely damage plants, limiting geographical distribution of natural populations and leading to major agronomical losses. Plants native to cold climates acquire increased freezing tolerance during exposure to low nonfreezing temperatures in a process termed cold acclimation. This involves many adaptative responses, including global changes in metabolite content and gene expression, and the accumulation of cold-regulated (COR) proteins, whose functions are largely unknown. Here we report that the chloroplast proteins COR15A and COR15B are necessary for full cold acclimation in Arabidopsis (Arabidopsis thaliana). They protect cell membranes, as indicated by electrolyte leakage and chlorophyll fluorescence measurements. Recombinant COR15 proteins stabilize lactate dehydrogenase during freezing in vitro. However, a transgenic approach shows that they have no influence on the stability of selected plastidic enzymes in vivo, although cold acclimation results in increased enzyme stability. This indicates that enzymes are stabilized by other mechanisms. Recombinant COR15 proteins are disordered in water, but fold into amphipathic a-helices at high osmolyte concentrations in the presence of membranes, a condition mimicking molecular crowding induced by dehydration during freezing. X-ray scattering experiments indicate protein-membrane interactions specifically under such crowding conditions. The COR15-membrane interactions lead to liposome stabilization during freezing. Collectively, our data demonstrate the requirement for COR15 accumulation for full cold acclimation of Arabidopsis. The function of these intrinsically disordered proteins is the stabilization of chloroplast membranes during freezing through a folding and binding mechanism, but not the stabilization of chloroplastic enzymes. This indicates a high functional specificity of these disordered plant proteins.

show abstract

A low-background-intensity focusing small-angle X-ray scattering undulator beamline

Cited by 482 publications

References 16 publications

Independent catalysis of the short form HisG from Lactococcus lactis

Independent catalysis of the short form HisG from Lactococcus lactis

First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer

Disordered Cold Regulated15 Proteins Protect Chloroplast Membranes during Freezing through Binding and Folding, But Do Not Stabilize Chloroplast Enzymes in Vivo

Contact Info

Product

Resources

About