Studies on stationary phase Vibrio sp. 2

Car, Nicholas George

Studies on stationary phase Vibrio sp. 2

Doctoral Thesis

1987

Publisher

University of Cape Town

Department

Department of Molecular and Cell Biology

Faculty

Faculty of Science

Abstract

Vibrio sp. 2 stationary phase cells are novel and interesting in that they are able to support phage growth in standing cultures, but not in shaken (aerated) cultures. Many physiological and morphological characteristics change when Vibrio sp. 2 stationary phase cells are removed from aeration: the relatively high levels of protein synthesis (Robb et al., 1977; 1978) decrease, with a concomitant increase in the levels of RNA synthesis; protein degradation rises from 1 %h⁻¹ to 2,9 %h⁻¹, and whilst the average cell length decreases, the range of cell lengths markedly increases. The magic spot nucleotides, ppGpp and pppGpp, which are present in stressed exponential phase Vibrio sp. 2 cells, are not detectable in stationary phase Vibrio cells. The specific proteolytic activity of shaking stationary phase cell-free extracts against the foreign protein [¹⁴C-me]globin was slightly higher than that of extracts from standing or exponential phase cells, while the specific proteolytic activity against [¹²⁵I]-insulin was slightly lower. On the basis of inhibitor studies and subcellular distribution, the proteolytic activities of the three types of extract differed. The addition of exogenous ATP to cell-free extracts either stimulated (Car & Woods, 1984) or depressed proteolytic activity depending on the procedure used to prepare the extracts. The proteolytic activity of fractions containing substantial amounts of membrane material, from all three types of extract, were markedly depressed by ATP. On preincubation of cell-free extracts from exponentially growing cells prior to assay of proteolytic activity, the activity was markedly stimulated (two- to four-fold). The stimulation,. however, varied, greatly between independently produced extracts. ATP had a much smaller stimulatory effect on preparations free of cell wall material from both types of stationary phase cells (the stimulation was less than two-fold), and the stimulation was not affected by preincubation of the extracts. Extracts prepared from starving cells, previously in exponential growth, were affected by the addition of ATP in a similar manner to that observed with stationary phase extracts (Car & Woods, 1984). Exponential and both types of stationary phase Vibrio sp. 2 cells have ATP-stimulated and ATP-depressed activities separable by ion-exchange chromatography, in addition to several other proteolytic activities. All types of Vibrio sp. 2 cells have a similar complement of proteolytic activities.