Resolution of protein disulphide-isomerase and glutathione-insulin transhydrogenase activities by covalent chromatography.

1. Protein disulphide-isomerase (EC 5.3.4.1) and glutathione-insulin transhydrogenase (EC 1.8.4.2) were resolved by covalent chromatography. Both activities, in a partially purified preparation from bovine liver, bind covalently as mixed disulphides to activated thiopropyl-Sepharose 6B, in a new stepwise elution procedure protein disulphide-isomerase is displaced in mildly reducing conditions whereas glutathione-insulin transhydrogenase is only displaced by more extreme reducing conditions. 2. This together with evidence for partial resolution of the two activities by ion-exchange chromatography, conclusively establishes that the two activities are not alternative activities of a single bovine liver enzyme. 3. Protein disulphide-isomerase, partially purified by a published procedure, has now been further purified by covalent chromatography and ion-exchange chromatography. The final material is 560-fold purified relative to a bovine liver homogenate; it has barely detectable glutathione-insulin transhydrogenase activity. 4. The purified protein disulphide-isomerase shows a single major band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis corresponding to a mol.wt. of 57000. 5. The purified protein disulphide-isomerase has Km values for 'scrambled' ribonuclease and dithiothreitol of 23 microgram/ml and 5.4 microM respectively and has a sharp pH optimum at 7.5. The enzyme has a broad thiol-specificity, and several monothiols, at 1mM, can replace dithiothreitol. 6. The purified protein disulphide-isomerase is completely inactivated after incubation with a 2-3 fold molar excess of iodoacetate. The enzyme is also significantly inhibited by low concentrations of Cd2+ ions. These findings strongly suggest the existence of a vicinal dithiol group essential for enzyme activity. 7. When a range of thiols were used as co-substrates for protein disulphide-isomerase activity, the activities were found to co-purify quantitatively, implying the presence of a single protein disulphide-isomerase of broad thiol-specificity. Glutathione-disulphide transhydrogenase activities, assayed with a range of disulphide compounds, did not co-purify quantitatively.     

COMPARATIVE STUDIES OF FLORAL MORPHOLOGY OF THE LABIATAE

Hillson , Charles J. (Pennsylvania State U., University Park.) Comparative studies of floral morphology of the Labiatae. Amer. Jour. Bot. 46(6): 451–459. Illus. 1959.—Comparative vascular studies of 39 species of mints from 27 genera reveal 2 basic stelar patterns: those in which the dorsal carpellary bundles are terminal in origin and those in which the dorsal carpellary bundles are basal in origin. Gradations of intermediate dorsal carpellary bundle divergence exist. Steles exhibiting terminal dorsal carpellary bundles are always associated with laminal ovules and are interpreted as being primitive. Marginal ovules are associated with floral steles exhibiting basal divergence of dorsal carpellary bundles and are regarded as being advanced. Adnation of traces seems to be a more reliable character in determining phylogenetic relationships than is connation. On the basis of 3 characters, viz: (1) position of dorsal carpellary bundle divergence, (2) ovule position and (3) degree of concrescence, a phylogenetic series of the 8 sub-families of Labiatae from advanced to primitive is proffered.     

THE VASCULAR TRANSITION REGION OF HELIANTHUS ANNUUS. I. BILATERAL AND UNILATERAL PATTERNS OF DIFFERENTIATION

The vascular transition region of Helianthus annuus L. may exhibit either a unilateral or a bilateral differentiation of primary phloem groups. The unilateral type of transition has been previously described but the bilateral type with its two variations is described here for the first time. The three patterns of transition are compared, and the significance of the variability of transition regions is discussed in terms of basic transition types and phylogenetic importance.     

THE VASCULAR TRANSITION REGION OF HELIANTHUS ANNUUS (COMPOSITAE). II. EFFECT OF IONIZING RADIATION

The vascular pattern in the transition region of Helianthus annuus L. may be altered by ionizing radiation in dosages ranging from 650–19,200 R. The alteration of the vascular cylinder is expressed by (1) the failure of normal tracheary elements to mature and (2) by the induction of accessory xylem and/or phloem strands. The anatomical alterations in the vasculature of the transition region are presented and their significance discussed in terms of seedling survival and the mechanisms controlling vascular differentiation.