Phosphate binding

P-glycoprotein (Pgp), a membrane pump often responsible for the multidrug resistance of cancer cells, undergoes conformational changes in the presence of substrates/modulators, or upon ATP depletion, reflected by its enhanced reactivity with the UIC2 monoclonal antibody. When the UIC2-shift was elicited by certain modulators (e.g. cyclosporin A or vinblastine, but not with verapamil or Tween 80), the subsequent binding of other monoclonal anti-Pgp Ig sharing epitopes with UIC2 (e.g. MM12.10) was abolished [Nagy, H., Goda, K., Arceci, R., Cianfriglia, M., ´ Mechetner, E. & Szabo Jr, G.

6p system191 01-06-2013 62 7   Download

We previously reported that GTS1 is involved in regulating ultradian oscillations of the glycolytic pathway induced by cyanide in cell suspensions as well as oscillations of energy metabolism in aerobic continuous cultures. Here, we screened a yeast cDNA library for proteins that bind to Gts1p using the yeast two-hybrid system and cloned multiple TDH cDNAs encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH).

10p research12 01-06-2013 41 3   Download

Cloning and over-expression of human glucose 6-phosphate dehydrogenase (Glc6P dehydrogenase) has for the first time allowed a detailed kinetic study of a preparation that is genetically homogeneous and in which all the protein molecules are of identical age. The steady-state kinetics of the recombinant enzyme, studied by fluorimetric initial-rate measurements, gave converging linear Lineweaver–Burk plots as expected for a ternary-complex mechanism.

8p research12 01-06-2013 66 4   Download

F1-ATPase is inactivated by entrapment of MgADP in catalytic sites and reactivatedbyMgATPorPi.Here, usinga mutant a3b3c complex of thermophilic F1-ATPase (aW463F/bY341W) and monitoring nucleotide binding by ¯uorescence quenching of an introduced tryptophan, we found that P i interfered with the binding of MgATP to F1-ATPase, but binding ofMgADPwas interferedwith to a lesser extent.

8p research12 29-04-2013 37 4   Download

T4DNA ligase is an Mg 2+ -dependent andATP-dependent enzyme that seals DNA nicks in three steps: it covalently binds AMP,transadenylates the nick phosphate,and cata-lyses formation of the phosphodiester bond releasing AMP.

11p tumor12 20-04-2013 45 7   Download

Spermadhesins are a family of 12–16 kDa proteins with a single CUB domain. PSP-I and PSP-II, the most abundant boar spermadhesins, are present in seminal plasma as a noncovalent heterodimer. Dimerization markedly affects the binding ability of the subunits. Notably, heparin and mannose 6-phosphate binding abilities of PSP-II are abolished, indicating that the corresponding binding sites may be located at (or near) the dimer interface.

8p fptmusic 12-04-2013 33 3   Download

The binding of inosine 5¢ phosphate (IMP) to ribonuclease A has been studied by kinetic and X-ray crystallographic experiments at high (1.5 A ˚ ) resolution. IMP is a competitive inhibitor of the enzyme with respect to Cp and binds to the catalytic cleft by anchoring three IMP molecules in a novel binding mode.

14p fptmusic 11-04-2013 59 3   Download

NMR and visible spectroscopy coupled to redox measurements were used to determine the equilibrium thermodynamic properties of the four haems in cytochrome c3 under conditions in which the protein was bound to lig-ands, the small anion phosphate and the protein rubredoxin with the iron in the active site replaced by zinc.

10p awards 06-04-2013 38 4   Download

2¢-Fluoro-2¢-deoxyuridine 3¢-phosphate (dU F MP) and arabinouridine 3¢-phosphate (araUMP) have non-natural furanose rings. dU F MP and araUMP were prepared by chemical synthesis and found to have three- to sevenfold higher affinity than uridine 3¢-phosphate (3¢-UMP) or 2¢-deoxy-uridine 3¢-phosphate (dUMP) for ribonuclease A (RNase A). These differ-ences probably arise (in part) from the phosphoryl groups of 3¢-UMP, dU F MP, and araUMP (pKa¼5.9) being more anionic than that of dUMP (pKa¼6.3). ...

12p awards 05-04-2013 29 2   Download

The extremely heat-stable 5¢-methylthioadenosine phos-phorylase from the hyperthermophilic archaeonPyrococcus furiosuswas cloned, expressed to high levels inEscherichia coli, and purified to homogeneity by heat precipitation and affinity chromatography. The recombinant enzyme was subjected to a kinetic analysis including initial velocity and product inhibition studies. The reaction follows an ordered Bi–Bi mechanism and phosphate binding precedes nucleo-side binding in the phosphorolytic direction....

11p awards 05-04-2013 39 2   Download

D-Ornithine aminomutase from Clostridium sticklandii comprises two strongly associating subunits, OraS and OraE, with molecular masses of 12 800 and 82 900 Da. Previous studies have shown that inEscherichia colithe recombinant OraS protein is synthesized in the soluble form and OraE as inclusion bodies. Refolding experiments also indicate that the interactions between OraS and OraE and the binding of either pyridoxal phosphate (PLP) or aden-osylcobalamin (AdoCbl) play important roles in the refolding process....

0p awards 05-04-2013 19 2   Download

In this report, we have focused our attention on identifying intracellular mammalian proteins that bind S100A12 in a Ca 2+ -dependent manner. Using S100A12 affinity chroma-tography, we have identified cytosolic NADP + -dependent isocitrate dehydrogenase (IDH), fructose-1,6-bisphosphate aldolase A (aldolase), glyceraldehyde-3-phosphate dehy-drogenese (GAPDH), annexin V, S100A9, and S100A12 itself as S100A12-binding proteins. Immunoprecipitation experiments indicated the formation of stable complexes between S100A12 and IDH, aldolase, GAPDH, annexin V and S100A9in vivo....

11p awards 05-04-2013 36 6   Download

We used a combined computer and biochemical approach to characterize human pyridoxine 5¢-phosphate oxidase (PNPO).The human PNPOgene is composed of seven exons and six introns, and spans approximately 8 kb.All exon/intron junctions contain the gt/ag consensus splicing site.The absence of TATA-like sequences, the presence of Sp1-binding sites and more importantly, the presence of CpG islands in the regulatory region of thePNPOgene are characteristic features of housekeeping genes.

10p dell39 03-04-2013 48 3   Download

Inorganic phosphate (Pi) and cofilin⁄actin depolymerizing factor proteins have opposite effects on actin filament structure and dynamics. Pi stabilizes the subdomain 2 in F-actin and decreases the critical concentration for actin polymerization. Conversely, cofilin enhances disorder in subdomain 2, increases the critical concentration, and accelerates actin treadmilling.

9p inspiron33 26-03-2013 57 4   Download

The guanine nucleotide-binding protein Ras occurs in solution in two different conformational states, state 1 and state 2 with an equilibrium constant K12 of 2.0, when the GTP analogue guanosine-5¢-(b,c-imido)tri-phosphate or guanosine-5¢-(b,c-methyleno)triphosphate is bound to the active centre.

15p galaxyss3 21-03-2013 39 3   Download

Sphingolipid activator proteins (SAPs), GM2 activator protein (GM2AP) and saposins (Saps) A–D are small, enzymatically inactive glycoproteins of the lysosome. Despite of their sequence homology, these lipid-binding and -transfer proteins show different specificities and varying modes of action. Water-soluble SAPs facilitate the degradation of membrane-bound glyco-sphingolipids with short oligosaccharide chains by exohydrolases at the membrane–water interface.

16p galaxyss3 19-03-2013 31 3   Download

Serine hydroxymethyltransferase (SHMT) fromBacillus stearothermophilus (bsSHMT) is a pyridoxal 5¢-phosphate-dependent enzyme that catalyses the conversion of l-serine and tetrahydrofolate to glycine and 5,10-methylene tetrahydrofolate. In addition, the enzyme catalyses the tetrahydrofolate-independent cleavage of 3-hydroxy amino acids and transamination.

14p galaxyss3 07-03-2013 49 4   Download

The invariant carboxylate residue which follows the Walker B motif (hyd4DE⁄D) in the nucleotide-binding domains (NBDs) of ATP-binding cassette transporters is thought to be involved in the hydrolysis of the c-phosphate of MgATP, either by activating the attacking water molecule or by promoting substrate-assisted catalysis.

13p galaxyss3 07-03-2013 30 5   Download

A hydrophilic cation-binding protein, PCaP1, was found to be stably bound to the plasma membrane inArabidopsis thaliana. PCaP1 was quanti-fied to account for 0.03–0.08% of the crude membrane fractions from roots and shoots.

16p media19 06-03-2013 39 3   Download

Possible binding proteins of CP12 in a green alga,Chlamydomonas rein-hardtii, were investigated. We covalently immobilized CP12 on a resin and then used it to trap CP12 partners. Thus, we found an association between CP12 and phosphoribulokinase (EC 2.7.1.19), glyceraldehyde 3-phosphate dehydrogenase (EC 1.2.1.13) and aldolase.

12p media19 06-03-2013 47 3   Download