Inhibition of riboflavin-sensitized photoinactivation of taka-amylase a by some inhibitors

Zusammenfassung Durch Gegenwart von Co⁺⁺ oder J^– oder Tryptophan oderp-Phenylendiamin wird die durch Riboflavin sensibilisierte Photoinaktivierung der Taka-Amylase A stark gehemmt. Die möglichen Mechanismen der Hemmungswirkung dieser Inhibitoren werden diskutiert.     

The amylolysis of a substituted starch substrate

Zusammenfassung Bei Anwendung eines Stärkesubstrates in Verbindung mit einem im Kohlenstoffatom 6 kovalent gebundenen Farbstoffmolekül gelingt die differentielle Messung der - und -Amylase Aktivität im Reaktions-medium.     

Inhibition of aggregation of heat-denatured Taka-amylase a by substrate

Zusammenfassung Das Substrat beschützt stark die Taka-Amylase A nicht nur gegen Wärme-Inaktivierung und -Denaturierung, sondern auch gegen Wärme-Aggregation. Diese Schutzwirkung resultiert aus der Stabilisierung der Protein-Struktur durch Bildung der Enzym-Substrat- und -Produkt-Komplexe.     

Evidence of G-protein-coupled receptor and substrate transporter heteromerization at a single molecule level

G-protein-coupled receptors (GPCRs) can constitute complexes with non-GPCR integral membrane proteins, while such interaction has not been demonstrated at a single molecule level so far. We here investigated the potential interaction between the thyrotropin receptor (TSHR) and the monocarboxylate transporter 8 (MCT8), a member of the major facilitator superfamily (MFS), using fluorescence cross-correlation spectroscopy (FCCS). Both the proteins are expressed endogenously on the basolateral plasma membrane of the thyrocytes and are involved in stimulation of thyroid hormone production and release. Indeed, we demonstrate strong interaction between both the proteins which causes a suppressed activation of G_q/11 by TSH-stimulated TSHR. Thus, we provide not only evidence for a novel interaction between the TSHR and MCT8, but could also prove this interaction on a single molecule level. Moreover, this interaction forces biased signaling at the TSHR. These results are of general interest for both the GPCR and the MFS research fields.     

Viscosity changes in a polymerized substrate due to the action of a hydrolytically depolymerizing enzyme

Zusammenfassung Nachweis einer Enzymbildung nach Azotobacter-Infektion durch Phagen mit Spezifität für Kapsel-Depolymerisierung. Das Verfahren erfasst die Enzymaktivität durch Registrierung der graduellen Veränderung der Viskosität des sich vorbereitenden Kapselstoffes.

---

---

Supported USPH Grant A102830.     

The unique substrate specificity of human AOC2, a semicarbazide-sensitive amine oxidase

Semicarbazide-sensitive amine oxidases (SSAOs) catalyze oxidative deamination of primary amines, but the true physiological function of these enzymes is still poorly understood. Here, we have studied the functional and structural characteristics of a human cell-surface SSAO, AOC2, which is homologous to the better characterized family member, AOC3. The preferred in vitro substrates of AOC2 were found to be 2-phenylethylamine, tryptamine and p-tyramine instead of methylamine and benzylamine, the favored substrates of AOC3. Molecular modeling suggested structural differences between AOC2 and AOC3, which provide AOC2 with the capability to use the larger monoamines as substrates. Even though AOC2 mRNA was expressed in many tissues, the only tissues with detectable AOC2-like enzyme activity were found in the eye. Characterization of AOC2 will help in evaluating the contribution of this enzyme to the pathological processes attributed to the SSAO activity and in designing specific inhibitors for the individual members of the SSAO family. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

2-Methoxybenzoyl phosphate: a new substrate for continuous fluorimetric and spectrophotometric acyl phosphatase assays

A new aromatic acyl phosphate, 2-methoxybenzoyl phosphate, has been synthesized. The compound shows an intrinsic fluorescence; it displays an intense emission band at 390 nm upon excitation in the near UV region. This band practically disappears after hydrolysis of the product. On the other hand, the product displays differences in the near UV absorption spectra measured before and after hydrolysis. The at 301 nm is 2720 M^–1 cm^–1, a value that is 4.3-fold higher than that of benzoyl phosphate (the usual substrate for acylphosphatase assay) at 283 nm. The main kinetic parameters of three different acylphosphatase molecular forms (the muscular isoenzyme and two subtypes of the organ common isoenzyme) were determined using both benzoyl phosphate and 2-methoxybenzoyl phosphate as substrates, and then compared. These kinetic data and the UV absorption and fluorescence properties of 2-methoxybenzoyl phosphate sugest that this compound has better substrate features than benzoyl phosphate, and can be used for both high sensitivity continuous fluorimetric and UV absorption spectrophotometric assays of acylphosphatase.     

Evidence for changes in cell shape from a 2-dimensional to a 3-dimensional substrate

Summary Chick embryo mesoderm cells were explanted to culture systems in vivo and in vitro and their subsequent movements were correlated with the external morphology as studied by SEM. In vitro cell movements are exaggerations of normal in vivo movements where a 2-dimensional substrate is encountered rather than a 3-dimensional environment.The authors are grateful to Mr J. Smith and Mrs S. Bulman who provided excellent technical assistance and to Mr G. L. C. McTurk who skillfully produced the scanning electron micrographs in the Leicester University Scanning Electron Microscope Unit.     

Evidence for changes in cell shape from a 2-dimensional to a 3-dimensional substrate.

Chick embryo mesoderm cells were explanted to culture systems in vivo and in vitro and their subsequent movements were correlated with the external morphology as studied by SEM. In vitro cell movements are exaggerations of normal in vivo movements where a 2-dimensional substrate is encountered rather than a 3-dimensional environment.     

d-Xylose,a substrate for the process of sugar active transport by the small intestine

Résumé Dans les cellules épithéliales de l'intestin grêle du Hamster, l'absorption de xylose comprend deux étapes: (1) uneentrée sensible à la phlorizine, et (2) une étape sensible au dinitrophénol, probablement équivalente à l'étape d'accumulation dépendant de l'énergie et typique des sucres actifs.

---

---

A preliminary account was presented at the VII Giornate Biochimiche Latine, S. Margherita Ligure (Genova 1963), p. 87.     

VLDL substrate properties and efficiency of their metabolic transformation by LPL

Summary A study was made of the regulation of the triglyceride hydrolysis catalysed by LPL from bovine milk, by the apoproteins from human plasma VLDL. Both isolated apolipoproteins, and those found on the surface of plasma VLDL particles, were investigated. A concentration-dependent activating action of apo C-II on the hydrolysis of emulsified triolein, and uncompetitive inhibition of VLDL triglyceride hydrolysis by apo C-III were found. It is suggested that VLDL lipolysis might be controlled in vivo trough the variation of the relative surface content of these enzymatic activity modulators.