Alfred S. Sussman

Role of Trehalose in Ascospores of Neurospora Tetrasperma

The anthrone-positive material extractable in 80 percent alcohol, whose disappearance is correlated with the breaking of dormancy, has been found to be a non-reducing sugar which yields only glucose upon hydrolysis. On the basis of its crystal structure, infrared spectrum, melting point, specific rotation, and chromatographic properties, this material has been identified as trehalose.

Purification and properties of trehalase(s) from Neurospora

Abstract Trehalase has been obtained in crystalline form from the mycelium of Neurospora crassa . During elution from the first pass through a DEAE-cellulose column, two peaks with trehalase activity were obtained. When the first of these was rechromatographed on DEAE-cellulose, two more major peaks were found. The enzymes of these fractions were compared and found to be similar in substrate specificity, response to inhibitors, pH optima, and Michaelis constants. However, small differences in the rate of inactivation of these enzymes at 50 ° were detected.

Effect of light and media upon growth and melanin formation in aureobasidium pullulans (de By.)Arn. (=pullularia pullulans)

Polysaccharides like dextrine and starch are shown to be the best carbon sources for the growth ofAureobasidium pullulans although growth is good upon a variety of other carbon sources. Light increases growth markedly when polysaccharides are the carbon source but not when other sugars are used. Variation in cell morphology is described in response to sugars and light. Extracellular granules, whose properties resemble those of melanin, are produced when dextrine is the carbon source in a defined medium containing asparagine as the source of nitrogen. The dark pigment was extracted from the walls of thick-walled brown cells ofA. pullulans and characterized as a melanin on the basis of several tests, including solubility and absorption spectrum.A. pullulans was grown on several defined and undefined media and the response of the fungus to light is shown to be determined by the medium, and the temperature at which the cultures are grown.

A comparison of the properties of two forms of tyrosinase from Neurospora crassa

The differences between a thermostable (Ts) and thermolabile (Tl) form of tyrosmase from Neurospora crassa have been studied by means of a spectrophotometric technique using ascorbate or ferrocyanide. No significant differences in substrate specificity have been found in a survey of over 30 substances which included mono-, di-, tri-, and conjugated phenols. Catechol was found to be a substrate, in contrast to previous findings. Michaelis-Menten constants for the two forms of enzyme acting on l-tyrosine and l- and d-hydroxyphenylalanine (Dopa) also were found to be similar, as were the pH optima on l-Dopa and phenyl-4-catechol, and the response to several inhibitors. On the other hand, the activity of the thermolabile enzyme was lost more rapidly than that of the thermostable one after incubation for 5–15 min. in high concentrations of urea and formamide. These data are interpreted as support for the conclusion that the two forms of the enzyme differ mainly in their secondary and tertiary structure and not in their active centers.

Accumulation of microfilaments in a colonial mutant of Neurospora crassa

A morphological mutant of Neurospora crassa, snowflake, is shown to contain filaments which are about 70 A in diameter, and up to several microns long, and which usually bunch in groups of a few to several hundred. They may be found longitudinally or transversely arranged with respect to the long axis of the cell and, in many cases, they run up to the plasma membrane, but not through it. The filaments often are arranged in crystalline arrays but may also be found as separate filaments. Sometimes the filaments are closely appressed to nuclei and may be found inside them. It is likely that the filaments are not the result of the dissociation of microtubules and are most likely microfilaments like those found in other organisms. Their relationship to the origin of certain morphological mutants in Neurospora is discussed.

A model for rhythmic and temperature-independent growth in ‘clock’ mutants of neurospora

The Q10 for the frequency (number of bands per 24 hours) of the ‘clock’ mutant (strain CL11A) ofNeurospora crassa over the range 20–30° C is close to 1.0. By contrast, that for the double mutant, ‘wrist watch’ (strain CL12a), is closer to 2 over this temperature range. Strain CL12a differs from ‘clock’ in other ways as well, including 1) decreased rate of linear extension and band size, 2) greater sensitivity of growth rate to high temperatures and, 3) masking of rhythmic growth below 15° C. The response to temperature of several colonial mutants and standard (‘wild-type’) strains was studied and it is shown that some strains are temperature-independent yet arhythmic. A temperature-compensation model is presented to explain the response of ‘clock’ mutants to temperature and it is concluded that they demonstrate a non-circadian free-running endogenous rhythm.

TYROSINASE AND THE RESPIRATION OF PUPAE OF PLATYSAMIA CECROPIA L.

1. The Qo2 of diapausing pupae of Platysamia cecropia is not affected by injection of substrates or inhibitors of the enzyme tyrosinase. 2. The Qo2 of homogenates prepared from such animals is markedly influenced by the above reagents. 3. On the basis of this and other data, it is concluded that tyrosinase does not function as a terminal oxidase in the respiration of the diapausing pupa.

Activation of Neurospora Ascospores by Organic Solvents and Furans

It has been shown previously (Emerson, 1948; Sussman, 1953) that dormant ascospores of Neurospora respond to very low concentrations of heterocyclic 5-membered ring compounds, including certain thiophene and furfural derivatives, and pyrrole. Although over 100 other substances were tried, only compounds with the above structure were active, with the exception of ethyl ether which was weakly effective in amounts 100-fold greater than were required when heterocyclics were used. Since that time, it was discovered that temperatures between 40 and 48? C, although insufficient to activate the ascospores, enhanced their sensitivity to chemical activators (Sussman, 1954a). Therefore, it seemed reasonable to expect that a more detailed investigation of the chemical specificity involved in the activation process might be possible by the use of this technique. Moreover, the observation that ethyl ether was also an activator suggested the possibility that other organic solvents might also serve in this way. That this surmise was correct is shown by the following experiments, which expand the list of chemical activators to include aliphatic alcohols, esters, and ketones, in addition to the furans used previously and in the present studies.

Antigenic Differences in the Surfaces of Hyphae and Rhizoids in Allomyces

Immunofluorescent techniques have demonstrated a difference in surface components of hyphae and rhizoids of Allomyces macrogynus. An antigenic component detected on the rhizoidal surface may be present, but masked, in the hyphal-wall matrix material. The system also allows visualization of the hyphal wall during aging, when changes from a smooth to a fissured surface are noted, and differences in adsorptive properties occur.

The development of tyrosinase and cytochrome oxidase activity in mutants of Glomerella cingulata

Abstract Cytochrome oxidase activity in the standard type and in four mutants of Glomerella cingulata has been shown to be constant throughout the mycelial development. This enzyme is concentrated on mitochondria-like particles which are extractable in hypertonic solutions of disaccharides. On the other hand, tyrosinase activity is concentrated in the supernatant and changes markedly during the development of the organism. This enzyme is present in the standard type and in two of the mutants, while the other two mutants have only minimal amounts. In the mycelia having such activity, tyrosinase increases precipitously, after the peak of growth has been reached, and then declines almost immediately, although small amounts of tyrosinase remain in the mycelium for as long as it is possible to obtain extracts. The evidence suggests that tyrosinase cannot function as the terminal oxidase during growth, although cytochrome oxidase can function in this way.