The lambda holin accumulates beyond the lethal triggering concentration under hyperexpression conditions.
Most bacteriophages terminate infection by creating lesions in the cytoplasmic membrane, which not only cause immediate cell death but also allow escape of a phage-encoded endolysin. Destruction of the peptidoglycan and cell lysis follows very rapidly, allowing efficient release of the progeny virions. These membrane lesions are formed by a small integral membrane protein called a holin. Holins have highly charged carboxyl-termini that are thought to have two transmembrane alpha-helical domains. Holins are believed to oligomerize and form large holes in the inner membrane. The prototype holin is the S protein from bacteriophage lambda. Scheduling of the lytic event is determined in part by the "structure directed initiation" or sdi translational control region. Inductions of S, cloned under a variety of native and nonnative promoters but with native translational control, resulted in cell lysis at about 1000 molecules of holin per cell, and thus do not produce biochemically useful amounts of S protein. By utilizing a plasmid-based system with the T7 RNA polymerase promoter in tandem with a consensus ribosome binding site, Coomassie blue-detectable quantities of S protein were obtained upon induction, corresponding to an approximately 100-fold increase over the normal lethal level of holin. Characterization of this expression system is presented and discussed with respect to the current model of holin function.
Smith
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Department of Biochemistry and Biophysics, Texas A&M University, College Station 77843, USA.
Chang
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CY
Young
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D000595:Amino Acid Sequence; D010582:Bacteriophage lambda / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity*; D001483:Base Sequence; D002463:Cell Membrane Permeability; D004279:DNA, Viral / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D004926:Escherichia coli; D015870:Gene Expression; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D014764:Viral Proteins / Q000096:biosynthesis* / Q000235:genetics
D004279:DNA, Viral; D008565:Membrane Proteins; C108624:S holin, bacteriophage lambda; D014764:Viral Proteins; C082001:bacteriophage lambda lysis effector protein S105; C082002:bacteriophage lambda lysis inhibitor protein S107; D010450:Endopeptidases; C001143:endolysin