A simple and reliable turbidimetric and kinetic assay for alpha-amylase that is readily applied to culture supernatants and cell extracts

Summary A simple, reliable and sensitive assay for alpha-amylase activity is reported, together with its theoretical derivation, that overcomes many of the problems encountered with other assays, especially when attempting to assay alpha-amylase activity in crude cell extracts or culture supernatants. The method relies on the reduction in turbidity that occurs upon digestion of a starch suspension with alpha-amylase. The initial rate of decrease in turbidity is shown to be proportional to a wide range of enzyme concentrations, permitting a rapid spectrophotometric and kinetic determination of alpha-amylase activity.     

Paraheliotropic leaf movement in Siratro as a protective mechanism against drought-induced damage to primary photosynthetic reactions: damage by excessive light and heat

Damage to primary photosynthetic reactions by drought, excess light and heat in leaves of Macroptilium atropurpureum Dc. cv. Siratro was assessed by measurements of chlorophyll fluorescence emission kinetics at 77 K (-196°C). Paraheliotropic leaf movement protected waterstressed Siratro leaves from damage by excess light (photoinhibition), by heat, and by the interactive effects of excess light and high leaf temperatures. When the leaves were restrained to a horizontal position, photoinhibition occurred and the degree of photoinhibitory damage increased with the time of exposure to high levels of solar radiation. Severe inhibition was followed by leaf death, but leaves gradually recovered from moderate damage. This drought-induced photoinhibitory damage seemed more closely related to low leaf water potential than to low leaf conductance. Exposure to leaf temperatures above 42°C caused damage to the photosynthetic system even in the dark and leaves died at 48°C. Between 42 and 48°C the degree of heat damage increased with the time of exposure, but recovery from moderate heat damage occurred over several days. The threshold temperature for direct heat damage increased with the growth temperature regime, but was unaffected by water-stress history or by current leaf water status. No direct heat damage occurred below 42°C, but in water-stressed plants photoinhibition increased with increasing leaf temperature in the range 31–42°C and with increasing photon flux density up to full sunglight values. Thus, water stress evidently predisposes the photosynthetic system to photoinhibition and high leaf temperature exacerbates this photoinhibitory damage. It seems probable that, under the climatic conditions where Siratro occurs in nature, but in the absence of paraheliotropic leaf movement, photoinhibitory damage would occur more frequently during drought than would direct heat damage.Abbreviations and symbols PFD photon flux area density - PSI, PSII photosyntem I, II - F _M, F _O, F _V maximum, instantaneous, variable fluorescence emission - PLM paraheliotropic leaf movement; all data of parameter of variation are mean ± standard error     

Physical and genetical characterisation of a second sex factor, SCP2, for Streptomyces coelicolor A3(2)

Summary Covalently closed circular (ccc) DNA of uniform monomer size (c. 18×10⁶ daltons) and restriction endonuclease cleavage pattern was isolated from strains of S. coelicolor A3(2) of differing constitution in respect of the SCP1 sex factor: SCP1⁺, SCP1, SCP1^- and NF (integrated SCP1). No such ccc DNA was found in strains of S. lividans 66 or S. parvulus ATCC 12434 whether or not they contained SCP1. These results confirmed that the 18×10⁶ dalton plasmid is not, and does not include, SCP1, which has not so far been isolated by any of a variety of methods.Genetic data served to identify a second sex factor, SCP2, postulated to be present in SCP2⁺ state in the starting strains and to be capable of mutation to a variant form, SCP2^*, with enhanced sex factor activity. From SCP2^* strains, SCP2^- cultures were isolated, at an average spontaneous frequency of about 0.8%. Crosses of pairs of SCP1^- SCP2^- strains were almost, but not completely, sterile; thus SCP1 and SCP2 probably contribute nearly all the fertility naturally occurring in the A3(2) strain. The two sex factors share the property of exerting an effect that may be comparable with lethal zygosis caused by F in E. coli: it is shown by SCP1-carrying strains against SCP1^-, or SCP2^* (but not SCP2⁺) strains against SCP2^- and is revealed as a narrow zone of growth inhibition surrounding the plasmid-carrying culture on a background of the appropriate plasmid-negative strain.Genetically defined SCP^2- strains lacked the ccc DNA found in SCP2⁺ and SCP2^* strains. Thus this DNA apparently represents the SCP2 sex factor. A preliminary restriction endonuclease cleavage map of SCP2 was constructed, with single sites for EcoRI and HindIII, four sites for SalPI (=PstI) and more than 20 sites for SalGI (SalI).     

Activation of α-1 adrenergic receptors modulates the control of left/right sidedness in rat embryos

Presomite stage rat embryos were cultured for 45–49 hr with medium containing various adrenergic agonists and antagonists. -Norepinephrine but not -norepinephrine (several orders of magnitude less potent than the -isomer at α-1 adrenergic receptors) resulted in a dose-dependent increase of situs inversus similar to that found for phenylephrine, an α-1 adrenergic agonist. Prazosin, an α-1 adrenergic antagonist, inhibited phenylephrine-induced situs inversus in a dose-dependent manner. Neither dexmedetomidine, an α-2 adrenergic agonist, nor isoproterenol, a β adrenergic agonist, caused situs inversus. These results provide pharmacological evidence that stimulation of α-1 but not of α-2 and β adrenergic receptors modulates the control of left/right sidedness in rat embryos.