The Oxidases of Gelasinospora tetrasperma in relation to the Endogenous Respiration

Abstract

SUMMARY:The active growth of Gelasinospora tetrasperma in a chemically defined liquid medium was completed in 5 days. Cytochrome was detected at all ages (spectroscopically and by manometric assay) and a cytochrome-cytochrome oxidase system was mainly responsible for 0, uptake. Phenol oxidase was not detected until the cultures had been growing 48 hr., and even then the proportion of the 0, uptake of the mycelium due to this enzyme was small, since light reversed carbon monoxide inhibition of the 0, uptake almost completely.Whereas the importance of cytochrome c oxidase as a terminal oxidase of plant respiration is now firmly established, the role, if any, of phenol oxidases in plant respiration is uncertain (James, 1 9 5 3~) . In the Basidiomycete Polystictus versicolor, which produced both cytochrome c oxidase and a phenol oxidase, laccase, only the former enzyme was involved in oxygen uptake (Boulter & Burges, 1955). The present work was undertaken to assess the relative importance of cytochrome c oxidase and the phenol oxidase, tyrosinase, in the endogenous respiration of Gelasinospora tetrasperma, an Ascomycete, which produces both these enzymes. METHODSCulture medium. Gelasinospora tetrasperma Dowding, obtained from Dr Dowding, was grown in a liquid medium of the following composition: D( + ) glucose, 10 g. ; asparagin, 5 g . ; KH,PO,, 0.5 g. ; Mg,SO,. 7H,O, 0.5 g . ; NaC1, 0-25 g.; biotin, 50pg.; thiamine, 1OOpg.; distilled water to 1 1. (glucose autoclaved separately ; thiamine sterilized by filtration).Mycelial suspensions from 48 hr. cultures, grown in 40 ml. medium at 25', were used for inoculations. Mycelium from two flasks was washed and transferred with 40 ml. water to the metal cup of an Atomix blender (M.S.E., London) and blended at full speed for 10 see. This agitation gave a fine mycelial suspension which was washed twice by centrifugation and resuspended in water; 1 ml. of suspension was used to inoculate each 250 ml. flask containing 90 ml. medium.Culture conditions. Cultures were incubated a t 25' on a reciprocal shaker (stroke 4 in., 75 oscillations/min.).Chemicals. Substrates and inhibitors were obtained from the British Drug Houses Ltd., and were of analytical grade when available. Acid and alkaline compounds were neutralized; unstable substrates were recrystallized and

Activation and Thermostability of Neurospora crassa Tyrosinase

“…Other reports have also suggested the existence of a 'protyrosinase' which can be activated by heating in Neurospora crassa (Horowitz & Shen, 1952;Schaeffer, 1953;Fox & Burnett, 1956) and in the ascomycete Getasinosporu tetrasperma (Boulter, 1957). Activation of tyrosinase and other copper oxidases can also be effected by various chemical agents, e.g., the grasshopper enzyme (Bodine et al 1944) is activated by urea and certain detergents, while ascorbic acid oxidase (Frieden & Maggiolo, 1957) is activated by a variety of substances (proteins, amino acids, thyroxine analogues, etc.)…”