@article{qiu_huber_booker_jain_leakey_fiscus_yau_ort_huber_2008, title={Increased protein carbonylation in leaves of Arabidopsis and soybean in response to elevated [CO2]}, volume={97}, ISSN={["1573-5079"]}, DOI={10.1007/s11120-008-9310-5}, abstractNote={While exposure of C3 plants to elevated [CO2] would be expected to reduce production of reactive oxygen species (ROS) in leaves because of reduced photorespiratory metabolism, results obtained in the present study suggest that exposure of plants to elevated [CO2] can result in increased oxidative stress. First, in Arabidopsis and soybean, leaf protein carbonylation, a marker of oxidative stress, was often increased when plants were exposed to elevated [CO2]. In soybean, increased carbonyl content was often associated with loss of leaf chlorophyll and reduced enhancement of leaf photosynthetic rate (Pn) by elevated [CO2]. Second, two-dimensional (2-DE) difference gel electrophoresis (DIGE) analysis of proteins extracted from leaves of soybean plants grown at elevated [CO2] or [O3] revealed that both treatments altered the abundance of a similar subset of proteins, consistent with the idea that both conditions may involve an oxidative stress. The 2-DE analysis of leaf proteins was facilitated by a novel and simple procedure to remove ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) from soluble soybean leaf extracts. Collectively, these findings add a new dimension to our understanding of global change biology and raise the possibility that oxidative signals can be an unexpected component of plant response to elevated [CO2].}, number={2}, journal={PHOTOSYNTHESIS RESEARCH}, author={Qiu, Quan-Sheng and Huber, Joan L. and Booker, Fitzgerald L. and Jain, Vanita and Leakey, Andrew D. B. and Fiscus, Edwin L. and Yau, Peter M. and Ort, Donald R. and Huber, Steven C.}, year={2008}, month={Aug}, pages={155–166} }
 @article{athwal_lombardo_huber_masters_fu_huber_2000, title={Modulation of 14-3-3 protein interactions with target polypeptides by physical and metabolic effectors}, volume={41}, DOI={10.1093/pcp/41.4.523}, abstractNote={The proteins commonly referred to as 14-3-3s have recently come to prominence in the study of protein:protein interactions, having been shown to act as allosteric or steric regulators and possibly scaffolds. The binding of 14-3-3 proteins to the regulatory phosphorylation site of nitrate reductase (NR) was studied in real-time by surface plasmon resonance, using primarily an immobilized synthetic phosphopeptide based on spinach NR-Ser543. Both plant and yeast 14-3-3 proteins were shown to bind the immobilized peptide ligand in a Mg2+-stimulated manner. Stimulation resulted from a reduction in KD and an increase in steady-state binding level (Req). As shown previously for plant 14-3-3s, fluorescent probes also indicated that yeast BMH2 interacted directly with cations, which bind and affect surface hydrophobicity. Binding of 14-3-3s to the phosphopeptide ligand occurred in the absence of divalent cations when the pH was reduced below neutral, and the basis for enhanced binding was a reduction in KD. At pH 7.5 (+Mg2+), AMP inhibited binding of plant 14-3-3s to the NR based peptide ligand. The binding of AMP to 14-3-3s was directly demonstrated by equilibrium dialysis (plant), and from the observation that recombinant plant 14-3-3s have a low, but detectable, AMP phosphatase activity.}, number={4}, journal={Plant and Cell Physiology}, author={Athwal, G. S. and Lombardo, C. R. and Huber, J. L. and Masters, S. C. and Fu, H. A. and Huber, S. C.}, year={2000}, pages={523–533} }
 @article{athwal_huber_huber_1998, title={Biological significance of divalent metal ion binding to 14-3-3 proteins in relationship to nitrate reductase inactivation}, volume={39}, ISSN={["0032-0781"]}, DOI={10.1093/oxfordjournals.pcp.a029303}, abstractNote={In this report we address two questions regarding the regulation of phosphorylated nitrate reductase (pNR; EC 1.6.6.1) by 14-3-3 proteins. The first concerns the requirement for millimolar concentrations of a divalent cation in order to form the inactive pNR:14-3-3 complex at pH 7.5. The second concerns the reduced requirement for divalent cations at pH 6.5. In answering these questions we highlight a possible general mechanism involved in the regulation of 14-3-3 binding to target proteins. We show that divalent cations (e.g. Ca2+, Mg2+ and Mn2+) bind directly to 14-3-3s, and as a result cause a conformational change, manifested as an increase in surface hydrophobicity. A similar change is also obtained by decreasing the pH from pH 7.5 to pH 6.5, in the absence of divalent cations, and we propose that protonation of amino acid residues brings about a similar effect to metal ion binding. A possible regulatory mechanism, where the 14-3-3 protein has to be "primed" prior to binding a target protein, is discussed.}, number={10}, journal={PLANT AND CELL PHYSIOLOGY}, author={Athwal, GS and Huber, JL and Huber, SC}, year={1998}, month={Oct}, pages={1065–1072} }
 @article{winter_huber_huber_1998, title={Identification of sucrose synthase as an actin-binding protein}, volume={430}, ISSN={["1873-3468"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-0032479338&partnerID=MN8TOARS}, DOI={10.1016/S0014-5793(98)00659-0}, abstractNote={Several lines of evidence indicate that sucrose synthase (SuSy) binds both G- and F-actin: (i) presence of SuSy in the Triton X-100-insoluble fraction of microsomal membranes (i.e. crude cytoskeleton fraction); (ii) co-immunoprecipitation of actin with anti-SuSy monoclonal antibodies; (iii) association of SuSy with in situ phalloidin-stabilized F-actin filaments; and (iv) direct binding to F-actin, polymerized in vitro. Aldolase, well known to interact with F-actin, interfered with binding of SuSy, suggesting that a common or overlapping binding site may be involved. We postulate that some of the soluble SuSy in the cytosol may be associated with the actin cytoskeleton in vivo.}, number={3}, journal={FEBS LETTERS}, author={Winter, H and Huber, JL and Huber, SC}, year={1998}, month={Jul}, pages={205–208} }
 @article{athwal_huber_huber_1998, title={Phosphorylated nitrate reductase and 14-3-3 proteins - Site of interaction, effects of ions, and evidence for an AMP-binding site on 14-3-3 proteins}, volume={118}, ISSN={["0032-0889"]}, DOI={10.1104/pp.118.3.1041}, abstractNote={The inactivation of phosphorylated nitrate reductase (NR) by the binding of 14-3-3 proteins is one of a very few unambiguous biological functions for 14-3-3 proteins. We report here that serine and threonine residues at the +6 to +8 positions, relative to the known regulatory binding site involving serine-543, are important in the interaction with GF14omega, a recombinant plant 14-3-3. Also shown is that an increase in ionic strength with KCl or inorganic phosphate, known physical effectors of NR activity, directly disrupts the binding of protein and peptide ligands to 14-3-3 proteins. Increased ionic strength attributable to KCl caused a change in conformation of GF14omega, resulting in reduced surface hydrophobicity, as visualized with a fluorescent probe. Similarly, it is shown that the 5' isomer of AMP was specifically able to disrupt the inactive phosphorylated NR:14-3-3 complex. Using the 5'-AMP fluorescent analog trinitrophenyl-AMP, we show that there is a probable AMP-binding site on GF14omega.}, number={3}, journal={PLANT PHYSIOLOGY}, author={Athwal, GS and Huber, JL and Huber, SC}, year={1998}, month={Nov}, pages={1041–1048} }
 @article{winter_huber_huber_1997, title={Membrane association of sucrose synthase: changes during the graviresponse and possible control by protein phosphorylation}, volume={420}, ISSN={["1873-3468"]}, url={http://www.scopus.com/inward/record.url?eid=2-s2.0-0031590792&partnerID=MN8TOARS}, DOI={10.1016/S0014-5793(97)01506-8}, abstractNote={Sucrose synthase (SuSy) plays an important role in sucrose degradation and occurs both as a soluble and as a membrane-associated enzyme in higher plants. We show that membrane association can vary in vivo in response to gravistimulation, apparently involving SuSy dephosphorylation, and is a reversible process in vitro. Phosphorylation of SuSy has little effect on its activity but decreases its surface hydrophobicity as reported with the fluorescent probe bis-ANS. We postulate that phosphorylation of SuSy (and perhaps other membrane proteins) is involved in the release of the membrane-bound enzyme in part as a result of decreased surface hydrophobicity.}, number={2-3}, journal={FEBS LETTERS}, author={Winter, H and Huber, JL and Huber, SC}, year={1997}, month={Dec}, pages={151–155} }
 @article{huber_redinbaugh_huber_campbell_1994, title={Regulation of maize leaf nitrate reductase activity involves both gene expression and protein phosphorylation}, volume={106}, DOI={10.1104/pp.106.4.1667}, abstractNote={Nitrate reductase (NR; EC 1.6.6.1) activity increased at the beginning of the photoperiod in mature green maize (Zea mays L.) leaves as a result of increased enzyme protein level and protein dephosphorylation. In vitro experiments suggested that phosphorylation of maize leaf NR affected sensitivity to Mg2+ inhibition, as shown previously in spinach. When excised leaves were fed 32P-labeled inorganic phosphate, NR was phosphorylated on seryl residues in both the light and dark. Tryptic peptide mapping of NR labeled in vivo indicated three major 32P-phosphopeptide fragments, and labeling of all three was reduced when leaves were illuminated. Maize leaf NR mRNA levels that were low at the end of the dark period peaked within 2 h in the light and decreased thereafter, and NR activity generally remained high. It appears that light signals, rather than an endogenous rhythm, account primarily for diurnal variations in NR mRNA levels. Overall, regulation of NR activity in mature maize leaves in response to light signals appears to involve control of gene expression, enzyme protein synthesis, and reversible protein phosphorylation.}, number={4}, journal={Plant Physiology}, author={Huber, J. L. and Redinbaugh, M. G. and Huber, S. C. and Campbell, W. H.}, year={1994}, pages={1667} }
 @article{huber_huber_campbell_redinbaugh_1992, title={REVERSIBLE LIGHT DARK MODULATION OF SPINACH LEAF NITRATE REDUCTASE-ACTIVITY INVOLVES PROTEIN-PHOSPHORYLATION}, volume={296}, ISSN={["0003-9861"]}, DOI={10.1016/0003-9861(92)90544-7}, abstractNote={Spinach ( Spinacia oleracea L.) leaf nitrate reductase (NADH:NR;NADH:nitrate oxidoreductase, EC 1.6.6.1) activity was found to rapidly change during light/dark transitions. The most rapid and dramatic changes were found in a form of NR which was sensitive to inhibition by millimolar concentrations of magnesium. This form of NR predominated in leaves in the dark, but was almost completely absent from leaves incubated in the light for only 30 min. When the leaves were returned to darkness, the NR rapidly became sensitive to Mg 2+ inhibition. Modulation of the overall reaction involving NADH as electron donor was also found when reduced methyl viologen was the donor (MV:NR), indicating that electron transfer had been blocked, at least in part, at or near the terminal molybdenum cofactor site. Changes in activity appear to be the result of a covalent modification that affects sensitivity of NR to inhibition by magnesium, and our results suggest that protein phosphorylation may be involved. NR was phosphorylated in vivo after feeding excised leave [ 32 P] P i . The NR subunit was labeled exclusively on seryl residues in both light and dark. Tryptic peptide mapping indicated three major 32 P-labeled phosphopeptide (Pp) fragments. Labeling of two of the P-peptides (designated Pp1 and 3) was generally correlated with NR activity assayed in the presence of Mg 2+ . In vivo , partial dephosphorylation of these sites (and activation of NR assayed with Mg 2+ ) occurred in response to light or feeding mannose in darkness. The light effect was blocked completely by feeding okadaic acid via the transpiration stream, indicating the involvement of type 1 and/or type 2A protein phosphatases in vivo . While more detailed analysis is required to establish a causal link between the phosphorylation status of NR and sensitivity to Mg 2+ inhibition, the current results are highly suggestive of one. Thus, in addition to the molecular genetic mechanisms regulating this key enzyme of nitrate assimilation, NR activity may be controlled in leaves by phosphorylation/ dephosphorylation of the enzyme protein resulting from metabolic changes taking place during light/dark transitions.}, number={1}, journal={ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS}, author={HUBER, JL and HUBER, SC and CAMPBELL, WH and REDINBAUGH, MG}, year={1992}, month={Jul}, pages={58–65} }
 @article{huber_huber_1992, title={Site-specific serine phosphorylation of spinach leaf sucrose-phosphate synthase}, volume={283}, number={3}, journal={Biochemical Journal (London, England : 1984)}, author={Huber, J. L. A. and Huber, S. C.}, year={1992}, pages={877} }
 @article{huber_hite_outlaw_huber_1991, title={INACTIVATION OF HIGHLY ACTIVATED SPINACH LEAF SUCROSE-PHOSPHATE SYNTHASE BY DEPHOSPHORYLATION}, volume={95}, ISSN={["0032-0889"]}, DOI={10.1104/pp.95.1.291}, abstractNote={Spinach (Spinacia oleracea L.) leaf sucrose-phosphate synthase (SPS) can be phosphorylated and inactivated in vitro with [γ-32P]ATP (JLA Huber, SC Huber, TH Nielsen [1989] Arch Biochem Biophys 270: 681-690). Thus, it was surprising to find that SPS, extracted from leaves fed mannose in the light to highly activate the enzyme, could be inactivated in an ATP-independent manner when desalted crude extracts were preincubated at 25°C before assay. The “spontaneous” inactivation involved a loss in activity measured with limiting substrate concentrations in the presence of the inhibitor, Pi, without affecting maximum catalytic activity. The spontaneous inactivation was unaffected by exogenous carrier proteins and protease inhibitors, but was inhibited by inorganic phosphate, fluoride, and molybdate, suggesting that a phosphatase may be involved. Okadaic acid, a potent inhibitor of mammalian type 1 and 2A protein phosphatases, had no effect up to 5 micromolar. Inactivation was stimulated about twofold by exogenous Mg2+ and was relatively insensitive to Ca2+ and to pH over the range pH 6.5 to 8.5. Radioactive phosphate incorporated into SPS during labeling of excised leaves with [32P]Pi (initially in the dark and then in the light with mannose) was lost with time when desalted crude extracts were incubated at 25°C, and the loss in radiolabel was substantially reduced by fluoride. These results provide direct evidence for action of an endogenous phosphatase(s) using SPS as substrate. We postulate that highly activated SPS contains phosphorylated residue(s) that increase activation state, and that spontaneous inactivation occurs by removal of these phosphate group(s). Inactivation of SPS in vivo caused by feeding uncouplers to darkened leaf tissue that had previously been fed mannose in the dark, may occur by this mechanism. However, there is no evidence that this mechanism is involved in light-dark regulation of SPS in vivo.}, number={1}, journal={PLANT PHYSIOLOGY}, author={HUBER, JL and HITE, DRC and OUTLAW, WH and HUBER, SC}, year={1991}, month={Jan}, pages={291–297} }
 @article{huber_pharr_huber_1990, title={PARTIAL-PURIFICATION AND CHARACTERIZATION OF STACHYOSE SYNTHASE IN LEAVES OF CUCUMIS-SATIVUS AND CUCUMIS-MELO - UTILIZATION OF A RAPID ASSAY FOR MYOINOSITOL}, volume={69}, ISSN={["0168-9452"]}, DOI={10.1016/0168-9452(90)90116-6}, abstractNote={A rapid enzymatic assay to quantitate myo-inositol using myo-inositol dehydrogenase-linked NAD reduction has been utilized to measure stachyose synthase activity Stachyose synthase activity in extracts of source leaves from C. sativus as well as sweet (cv. Noy Yizre'el) and non-sweet (cv. Birdsnest) genotypes of C. melo was separated completely from galactosidase and galactohydrolase activity via MONO Q anion exchange chromatography. As a result, it was determined for the first time that galactohydrolase activity present in leaf extracts is inhibited in the presence of raffinose and does not interfere with accurate estimation of stachyose synthase activity. The apparent Km values for stachyose synthase for galactinol were determined to be 4.6 mM, 2.4 mM and 3.0 mM for C. sativus, C. melo cv. Birdsnest and cv. Noy Yizer'el, respectively. The apparent Km value for raffinose was estimated to be approximately 15 mM for both the cucumber and melon source tissues. No in vitro metabolite effectors of the partially purified stachyose synthase have been identified. Thus, in vitro regulatory and kinetic properties of source leaf stachyose synthase are similar between species or cultivars of Cucumis which are predicted to differ significantly in sink carbohydrate metabolism.}, number={2}, journal={PLANT SCIENCE}, author={HUBER, JLA and PHARR, DM and HUBER, SC}, year={1990}, pages={179–188} }
 @article{huber_huber_nielsen_1989, title={PROTEIN-PHOSPHORYLATION AS A MECHANISM FOR REGULATION OF SPINACH LEAF SUCROSE-PHOSPHATE SYNTHASE ACTIVITY}, volume={270}, ISSN={["0003-9861"]}, DOI={10.1016/0003-9861(89)90551-1}, abstractNote={Studies were conducted to determine whether protein phosphorylation may be a mechanism for regulation of spinach (Spinacia oleracea L.) leaf sucrose-phosphate synthase (SPS), shown previously to be light-dark regulated by some type of covalent modification. Radioactive phosphate was incorporated into the 120-kDa subunit of SPS during labeling of excised leaves with [32P]Pi, as shown by immunoprecipitation and denaturing gel electrophoresis of the enzyme. Conditions which activated the enzyme (illumination of leaves or mannose treatment of leaf discs in darkness) reduced the incorporation of radiolabel into SPS in the in vivo system. The partially purified SPS protein could also be phosphorylated in vitro using [γ-32P]ATP. In the in vitro system, the incorporation of radiolabel into the 120-kDa subunit of SPS was dependent on time and magnesium concentration, and was closely paralleled by inactivation of the enzyme. These results provide the first evidence to establish protein phosphorylation as a mechanism for the covalent regulation of SPS activity.}, number={2}, journal={ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS}, author={HUBER, JLA and HUBER, SC and NIELSEN, TH}, year={1989}, month={May}, pages={681–690} }