Kinetic measurements of the interaction between an oligosaccharide and various lectins were performed using a biosensor based on surface plasmon resonance (SPR). A glycopeptide, prepared from asialofetuin and having a nearly homogeneous N-linked sugar chain, was immobilized on the surface of a sensor chip via the amino groups of its peptide moiety. The interactions of this bound glycopeptide with six lectins [Sambucus sieboldiana lectin, Maackia amurensis lectin, Aleuria uuruntia lectin, Ricinus communis agglutinin-1 20 (RCA,,,), Datura stramonium lectin (DSA) and Phaseolus vulgaris leukoagglutinating lectin] were monitored in real-time with the change in the SPR response. Of these lectins, only RCA,,, and DSA showed an increase in the SPR response, indicating that these two lectins bound specifically to the immobilized glycopeptide. The other lectins did not show any significant changes in the SPR response. These results are in good agreement with the binding specificity previously demonstrated with affinity chromatography. The association-rate constant (k,,,) and the dissociation-rate constant (k,,,,) for the glycopeptide-RCA,,, interaction were 3.4X lo5 M-' s-' and 2.1 X lop3 s-', respectively. The k,,, and kdlSs determined for DSA were S.7X105 M-' s-' and 1.3X 10-3 sp', respectively. Furthermore, the relative binding molar ratio to the glycopeptide was three times higher for RCA,,, than for DSA, suggesting that this sugar chain possesses three binding sites for RCA,,, and one for DSA. These parameters are expected to provide useful information for defining the interaction between oligosaccharides and lectins.Many aspects of the interactions between lectins and glycoproteins have been studied, especially regarding the recognition of molecules. These interactions have been used for the detection of the oligosaccharides of glycoproteins. In conventional methods, the recognition and adsorption properties of lectins with the specific sugar residues of glycoproteins are used for chromatographic purification and electrophoretic analytical techniques [l-31. However, it is not easy to elucidate the reaction mechanisms. Furthermore, for kinetic studies of the interactions between lectins and glycoproteins, it is usually necessary to label the molecules, either lectins or carbohydrates [4, 51. This has been a time-consuming step in these studies. The method introduced here requires no labeling, and the interaction phenomena of biological molecules can be monitored directly by the surface plasmon resonance (SPR).The phenomena of SPR was studied by Otto [6] and Kretschman and Raether [7], and it was used as a chemical Correspondence to Y. Shinohara,

An a-amylase[ƒ¿-1,4-glucan 4-glucanohydrolase, EC 3.2.1.1.], found in the culture filtrate of a strain of Thermoactinomyces vulgaris, was purified by ammonium sulfate fractionation , and DEAF-cellulose and CM-cellulose chromatographies. The purified enzyme showed a single band on disc gel electrophoresis. The optimum reaction pH and temperature were determined to be around pH 5.0 and 70°C. The isoelectric point was determined to be pH 5.2. The a-amylase was stabilized by Cal+. The a-amylase was found to hydrolyze pullulan to panose. Therefore, the hydrolytic pattern of this enzyme is different from those of pullulanase and isopullulanase. The authors have screened thermophilic acti nomycetes strains for a-amylase production and found that one actinomycetes strain iso lated from a soil sample at 50°C produced a new type of ƒ¿-amylase. The a-amylase had the ability to hydrolyze pullulan and showed different properties from the Thermoactino myces a-amylase reported by M. J. Kuo and P. A. Hartman.1,2) Since no ƒ¿-amylases have been reported to hydrolyze the part of pullulan consisting of maltotriose unit, this enzyme seems to be a new type of a-amylase. The present report describes the procedure of puri fication, some properties, and the pattern of pullulan hydrolysis of this ƒ¿-amylase. MATERIALS AND METHODS Microorganism and culture. A thermophilic acti nomycetes strain showing both ƒ¿-amylase and pullulan hydrolyzing activities was isolated from soil at 50°C.

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a b s t r a c tHvLhcb1 a major light-harvesting chlorophyll a/b-binding protein in barley, is a critical player in sustainable growth under Fe deficiency. Here, we demonstrate that Fe deficiency induces phosphorylation of HvLhcb1 proteins leading to their migration from grana stacks to stroma thylakoid membranes. HvLhcb1 remained phosphorylated even in the dark and apparently independently of state transition, which represents a mechanism for short-term acclimation. Our data suggest that the constitutive phosphorylation-triggered translocation of HvLhcb1 under Fe deficiency contributes to optimization of the excitation balance between photosystem II and photosystem I, the latter of which is a main target of Fe deficiency.

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