The influence of truncating the carboxy-terminal amino acid residues of streptococcal enolase on its ability to interact with canine plasminogen.

The native octameric structure of streptococcal enolase from Streptococcus pyogenes increasingly dissociates as amino acid residues are removed one by one from the carboxy-terminus. These truncations gradually convert native octameric enolase into monomers and oligomers. In this work, we investigated how these truncations influence the interaction between Streptococcal enolase and canine plasminogen.

The Energetics of Streptococcal Enolase Octamer Formation: The Quantitative Contributions of the Last Eight Amino Acids at the Carboxy-Terminus.

The enolase produced by Streptococcus pyogenes is a homo-octamer whose overall shape resembles that of a donut. The octamer is best described as a tetramer of dimers. As such, it contains two types of interfaces.

A multilaboratory comparison of calibration accuracy and the performance of external references in analytical ultracentrifugation.

Analytical ultracentrifugation (AUC) is a first principles based method to determine absolute sedimentation coefficients and buoyant molar masses of macromolecules and their complexes, reporting on their size and shape in free solution. The purpose of this multi-laboratory study was to establish the precision and accuracy of basic data dimensions in AUC and validate previously proposed calibration techniques. Three kits of AUC cell assemblies containing radial and temperature calibration tools and a bovine serum albumin (BSA) reference sample were shared among 67 laboratories, generating 129 comprehensive data sets.

tRNAGlu increases the affinity of glutamyl-tRNA synthetase for its inhibitor glutamyl-sulfamoyl-adenosine, an analogue of the aminoacylation reaction intermediate glutamyl-AMP: mechanistic and evolutionary implications.

For tRNA-dependent protein biosynthesis, amino acids are first activated by aminoacyl-tRNA synthetases (aaRSs) yielding the reaction intermediates aminoacyl-AMP (aa-AMP). Stable analogues of aa-AMP, such as aminoacyl-sulfamoyl-adenosines, inhibit their cognate aaRSs. Glutamyl-sulfamoyl-adenosine (Glu-AMS) is the best known inhibitor of Escherichia coli glutamyl-tRNA synthetase (GluRS).

The interaction of streptococcal enolase with canine plasminogen: the role of surfaces in complex formation.

The enolase from Streptococcus pyogenes (Str enolase F137L/E363G) is a homo-octamer shaped like a donut. Plasminogen (Pgn) is a monomeric protein composed of seven discrete separated domains organized into a lock washer. The enolase is known to bind Pgn.

eIF4E3 acts as a tumor suppressor by utilizing an atypical mode of methyl-7-guanosine cap recognition.

Recognition of the methyl-7-guanosine (m(7)G) cap structure on mRNA is an essential feature of mRNA metabolism and thus gene expression. Eukaryotic translation initiation factor 4E (eIF4E) promotes translation, mRNA export, proliferation, and oncogenic transformation dependent on this cap-binding activity. eIF4E-cap recognition is mediated via complementary charge interactions of the positively charged m(7)G cap between the negative π-electron clouds from two aromatic residues.

The interaction of canine plasminogen with Streptococcus pyogenes enolase: they bind to one another but what is the nature of the structures involved?

For years it has been clear that plasminogen from different sources and enolase from different sources interact strongly. What is less clear is the nature of the structures required for them to interact. This work examines the interaction between canine plasminogen (dPgn) and Streptococcus pyogenes enolase (Str enolase) using analytical ultracentrifugation (AUC), surface plasmon resonance (SPR), fluorescence polarization, dynamic light scattering (DLS), isothermal titration calorimetry (ITC), and simple pull-down reactions.

Structure-function relationships of wheat flavone O-methyltransferase: Homology modeling and site-directed mutagenesis.

Background: Wheat (Triticum aestivum L.) O-methyltransferase (TaOMT2) catalyzes the sequential methylation of the flavone, tricetin, to its 3'-methyl- (selgin), 3',5'-dimethyl- (tricin) and 3',4',5'-trimethyl ether derivatives. Tricin, a potential multifunctional nutraceutical, is the major enzyme reaction product.

Two fundamentally distinct PCNA interaction peptides contribute to chromatin assembly factor 1 function.

Chromatin assembly factor 1 (CAF-1) deposits histones H3 and H4 rapidly behind replication forks through an interaction with the proliferating cell nuclear antigen (PCNA), a DNA polymerase processivity factor that also binds to a number of replication enzymes and other proteins that act on nascent DNA. The mechanisms that enable CAF-1 and other PCNA-binding proteins to function harmoniously at the replication fork are poorly understood. Here we report that the large subunit of human CAF-1 (p150) contains two distinct PCNA interaction peptides (PIPs).

Reduction of canine plasminogen leads to an expanded molecule which precipitates.

Canine plasminogen is made up of seven domains. In each domain there are several cysteines that are linked by disulfide bonds. Reduction of a limited number of the cystines destabilizes the protein such that it precipitates.

Structure-activity relationships of wheat flavone O-methyltransferase: a homodimer of convenience.

Wheat flavone O-methyltransferase catalyzes three sequential methylations of the flavone tricetin. Like other flavonoid O-methyltransferases, the protein is a homodimer. We demonstrate, using analytical ultracentrifugation, that perchlorate dissociates the dimer into monomers.

Canine plasminogen: spectral responses to changes in 6-aminohexanoate and temperature.

We studied the near UV absorption spectrum of canine plasminogen. There are 19 tryptophans, 19 phenylalanines and 34 tyrosines in the protein. 4th derivative spectra optimized for either tryptophan or tyrosine give a measure of the polarity of the environments of these two aromatic amino acids.

Influence of temperature on the conformation of canine plasminogen: an analytical ultracentrifugation and dynamic light scattering study.

Plasminogen is known to undergo an extremely large conformational change when it binds ligands; the two well-established conformations are either closed (absence of external ligand) or open (presence of external ligand). We show here that plasminogen is more complicated than can be accommodated by a two-state, closed/open, model. Temperature changes induce large structural changes which can be detected with either dynamic light scattering or analytical ultracentrifugation.

Interactions between canine plasminogen and 8-anilino-1-naphthalene sulfonate: structural insights from a fluorescent probe.

Canine plasminogen (DPgn) binds the fluorescent molecule, 8-anilino-1-naphthalene sulfonate (ANS). Binding is favored by low pH as shown by steady-state fluorescence, native PAGE and isothermal titration calorimetry. Binding occurs at multiple sites.

The powerful high pressure tool for protein conformational studies.

The pressure behavior of proteins may be summarized as a the pressure-induced disordering of their structures. This thermodynamic parameter has effects on proteins that are similar but not identical to those induced by temperature, the other thermodynamic parameter. Of particular importance are the intermolecular interactions that follow partial protein unfolding and that give rise to the formation of fibrils.

Characterization of a complex formed between human plasminogen and recombinant sheep prion: pressure and thermal sensitivity of complex formation.

Scrapie is thought to be caused by one or more conformations of a proteinacious particle called a prion. The infectious form(s) is referred to as the scrapie form of the prion protein (PrPsc) whereas a benign form, the cellular conformer, is referred to as PrPC. The cellular conformation of the sheep prion protein formed a 1:1 complex with human plasminogen.

Role of Serine 286 in cosubstrate binding and catalysis of a flavonol O-methyltransferase.

O-Methyltransferases catalyze the transfer of the methyl groups of S-adenosyl-L-methionine to specific hydroxyl groups of several classes of flavonoid compounds. Of the several cDNA clones isolated from a Chrysosplenium americanum library, FOMT3' encodes the 3'/5'-O-methylation of partially methylated flavonols. The recombinant protein of another clone, FOMTx which differs from FOMT3' by a single amino acid residue (Ser286Arg) exhibits no enzymatic activity towards any of the flavonoid substrates tested.

The fate of the prion protein in the prion/plasminogen complex.

The cellular prion protein (PrP(c)) forms complexes with plasminogen. Here, we show that the PrP(c) in this complex is cleaved to yield fragments of PrP(c). The cleavage is accelerated by plasmin but does not appear to be dependent on it.

The influence of temperature and osmolyte on the catalytic cycle of cytochrome c oxidase.

The influence of temperature on cytochrome c oxidase (CCO) catalytic activity was studied in the temperature range 240-308 K. Temperatures below 273 K required the inclusion of the osmolyte ethylene glycol. For steady-state activity between 278 and 308 K the activation energy was 12 kcal x mol-1; the molecular activity or turnover number was 12 s-1 at 280 K in the absence of ethylene glycol.

The effects of osmotic and hydrostatic pressures on macromolecular systems.

Osmotic pressure and hydrostatic pressure can be used effectively to probe the behavior of biologically important macromolecules and their complexes. Using the two techniques requires a theoretical framework as well as knowledge of the more common pitfalls. Both are discussed in this review in the context of several examples.

Water as it applies to the function of enzymes.

Escherichia coli and Saccharomyces cerevisiae can metabolize, grow, and divide over osmotic pressures ranging from 0.24 atm to about 100 atm [Record, T. M.

Lactation alters gamma-aminobutyric acid neuronal activity in the hypothalamus and cerebral cortex in the rat.

Gamma-aminobutyric acid (GABA) neurons terminating in the hypothalamus have been implicated in the neuroendocrine regulation of reproductive hormones, particularly luteinizing hormone (LH) and prolactin. The aim of this study was to examine whether GABAergic neuronal activity in the hypothalamus was modified during lactation, and whether any observed changes correlated with changes in secretion of these hormones. Animals were divided into three experimental groups: diestrous controls, lactating with pups present (with pups), and lactating with pups removed for 4 h (without pups).

Reaction of canine plasminogen with 6-aminohexanoate: a thermodynamic study combining fluorescence, circular dichroism, and isothermal titration calorimetry.

The thermodynamics of the binding of 6-aminohexanoate (6-AH) to dog glu-plasminogen has been studied. Fluorescence titrations revealed four binding sites. Three yielded positive fluorescence changes on ligand binding; one yielded a negative fluorescence change.

Understanding the fluorescence changes of human plasminogen when it binds the ligand, 6-aminohexanoate: a synthesis.

This work attempts to explain several aspects of the response of plasminogen to 6-aminohexanoate (6-AH). These responses include the overall fluorescent changes that occur when plasminogen binds the ligand, the changes shown by the individual domains when they bind the ligand, and the changes in structure shown by the holoprotein when it binds 6-AH. The results have implications for understanding the physicochemical behavior of all kringle based proteins.