Cytochemical Localization of Certain Phosphatases in Escherichia coli

Abstract

Cytochemical studies of Escherichia coli at the light and electron microscopic levels have revealed alkaline phosphatase, hexose monophosphatase, and cyclic phosphodiesterase reaction products in the periplasmic space and at the cell surface. In preparations for both light and electron microscopy, reaction product filled polar caplike enlargements of the periplasmic space, such as those described in plasmolyzed cells, indicating significant terminal concentrations of these enzymes; dense substance was often seen within these polar caps in morphological specimens. Staining of the bacterial surface was commonly encountered, but could represent artifactual accumulation of precipitate along the cell wall. Alkaline phosphatase was demonstrated with several substrates (ethanolamine phosphate, glycerophosphate, p -nitrophenylphosphate, and glucose-6-phosphate) over a wide p H range in a bacterial strain (C-90) known to be constitutive for this enzyme, whereas strains deficient in this enzyme (U-7, repressed K-37), showed no activity with these substrates. Hexose monophosphatase and cyclic phosphodiesterase activities were characterized by reaction-product deposition with specific substrates at acid or neutral, but not at alkaline, p H in strains of E. coli lacking alkaline phosphatase (U-7 and repressed K-37). Fixation in Formalin or the use of calcium as a capture reagent seemed to interfere with periplasmic staining in cells prepared for electron microscopy. Formalin fixation had little effect on biochemical assays of the phosphatase activity of intact cells in suspension, but partially reduced the activity evident in sonically treated extracts or in suspensions of dispersed cryostat sections. Glutaraldehyde treatment impaired enzyme activity more drastically.

Antimicrobial Activity, Uses and Mechanism of Action of Glutaraldehyde

“…Ellar et al (1971) found that glutaraldehyde prevented the selective release of certain enzymes from the cytopl&mic membrane of Micrococcus lysodeikticus. Various concentrations of glutaraldehyde inactivate several periplasmic enzymes (Done et al 1965;Wang & Tu 1969;Wetzel et al 1970), including APase (Wetzel et a! .…”

Sequential Degranulation of the Two Types of Polymorphonuclear Leukocyte Granules During Phagocytosis of Microorganisms

“…In addition, reaction product for this enzyme has been localized at the fine-structural level in the periplasmic space in other strains of E . coli (26) . taining reaction product for alkaline phosphatase (Fig.…”

Tsr–GFP accumulates linearly with time at cell poles, and can be used to differentiate ‘old’ versus ‘new’ poles, in Escherichia coli

“…Cluster assembly appears to be a stochastic process involving competition between nucleation of new clusters and growth of existing clusters (Thiem et al, 2007;Thiem and Sourjik, 2008). Similarly, secreted proteins such as alkaline phosphatase or maltose-binding protein collect differentially in the periplasmic spaces of the poles, a phenomenon known as formation of 'polar caps' (Wetzel et al, 1970;Dietzel et al, 1978).…”