A bacteriophage-induced depolymerase active on Klebsiella K11 capsular polysaccharide

Bessler, Wolfgang G.; Freund-Mölbert, E; Knüfermann, H.; Rudolph, Claus; Thurow, Horst; Stirm, Stephan

doi:10.1016/0042-6822(73)90293-6

Cited by 54 publications

(43 citation statements)

References 36 publications

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“…In fact, Yurewicz et al [24] and Bessler et a/. [25] strongly suggested this possibility in their reports on Aerohacter and Klebsiella phages. Several studies concerning the enzymic and molecular properties of this kind of proteins show, however, that they consist of several nonidentical subunits, in contrast to the base-plate parts of the Salmonello phages P22, &15 [2] and E~~ [4].…”

Section: Dlscussionmentioning

confidence: 88%

Enzymic and Molecular Properties of Base-Plate Parts of Bacteriophage P22

Iwashita

Kanegasaki

1976

Eur J Biochem

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Using 14C-labeled Salmonella bacterial cells as the substrate, the enzymic and molecular properties of the base-plate parts of phage P22 were studied. The base-plate part consisted of a single protein species which cleaved extensively the 0-antigen of Salmonella typhimurium, Salmonelly schottmuellerie and with somewhat slomer rate that of Salmonella typhi, releasing oligosaccharide products with rhamnose at the reducing end. Much less cleavage was observed with a strain of S. typhimurium lysogenic for P22, and no significant reaction with Salmonella anatum, Salmonella newington and Salmonella minneapolis.The base-plate part enzyme was a very heat-stable protein and only 10-20 % loss was observed after treatment at 85 "C for 5 min. The pH optimum of the enzyme was around 7.5, and the glycosidase activity was not influenced by the ionic strength (25 -250 mM) of the medium or the presence of Mg2The molecular weight of the base-plate part was 320 000 by sedimentation equilibrium. Dodecylsulphate-acrylamide gel electrophoresis revealed a single band of molecular weight 77 000, indicating that a single base-plate part corresponds to a tetramer of identical subunits.Circular dichroism spectra of P22 base-plate parts showed a major contribution of j structure. The protein was rich in acidic amino acids, glycine and serine.It is now certain that the base-plate parts of at least some smooth-specific Salmonefla phages consist of enzymes with various activities which require the specific structure of 0-antigen polysaccharide to act on [l-41. The 0-antigen is the peripheral poration of the lipopolysaccharide, an important constituent of the outer membrane of gram-negative organisms [5].The base-plate parts of phages P22, &15 and c34 are specific endoglycosidases and cleave the host 0-antigen, which is composed of a linear repeating sequence of mannosyl-rhamnosyl-galactose. With phage P22 [l] and E~~ [2], the cleavage is caused by the specific breakage between rhamnose and galactose, and with E~~ [3, 41 the cleavage sites are galactosylmannose linkages. The base-plate, composed of the base-plate parts, is essential for phage infection [3,6]. Although information concerning the molecular properties of these enzyme proteins and their mode of action are required for the understanding of the molecular interaction between phage and host bacterium during the early stage of phage-infection process, such information has not been available so far. As reported earlier, all these base-plate parts can be obtained from phage lysates [7]. However, we have chosen a mutant of P22 in this report in order to obtain a highly purified preparation of base-plate parts. This phage mutant can not produce head particles in nonpermissive hosts, because of the defect of head gene 5 [8,9]. This paper describes the enzymic and molecular properties of the base-plate parts of P22, obtained in this manner. MATERIALS AND METHODS Bac terial Strains and PhagesThe bacteria used in this experiment were Salmonella typhimurium strains DB 21, DB 74 (permissible hos...

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Section: Dlscussionmentioning

confidence: 88%

Enzymic and Molecular Properties of Base-Plate Parts of Bacteriophage P22

Iwashita

Kanegasaki

1976

Eur J Biochem

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“…25 was isolated from Freiburg sewage. It forms plaques with acapsular halos (2,40) on Klebsiella 2002/49.…”

Section: Methodsmentioning

confidence: 99%

“…4. It is also worth noting that the unaltered amount of Glc in oligosaccharide P1 shows that the phage enzyme acts by hydrolysis and not by elimination and that -analogous to similar cases (2,40) -the spike-shaped organelles occasionally visible at the tail tips of phage 25 particles ( Fig. 1) may constitute the depolymerase.…”

Section: Vol 130 1977mentioning

confidence: 99%

“…25-isolation, propagation, purification, and electron microscopy. UsingKlebsiella 2002/49 as a prospective host, phage 25 was enriched from Freiburg sewage (39) and selected because of its plaque morphology, i.e., formation of acapsular haloes (2,40). The virus was purified by 10 successive single-plaque isolations.…”

Section: Vol 130 1977mentioning

confidence: 99%

“…25 (Fig. 3), a virus that forms plaques with acapsular haloes, which was selected for this property (2,40,42). The smallest K25 oligosaccharide obtained by exhaustive depolymerization (P1) is a tetrasaccharide ending in reducing Gal.…”

Section: Vol 130 1977mentioning

confidence: 99%

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