PMID:6363718

Citation	Strauss, H and King, J (1984) Steps in the stabilization of newly packaged DNA during phage P22 morphogenesis. J. Mol. Biol. 172:523-43
Abstract	The protein products of three adjacent P22 genes, 4, 10 and 26, are required for the stabilization of DNA newly packaged into P22 phage capsids. We have isolated unstable DNA containing capsids from cells infected with mutants defective in these genes. All three classes could be converted into mature phage in vitro, confirming that they represent intermediates in particle maturation. The first of the three proteins to add to the newly filled capsids is gp4, followed by gp10 and gp26. The active form of gp4 sediments at 3 S, while the active forms of both gp10 and gp26 sediment at 5 S. These soluble subunits appear to polymerize onto the newly filled capsids to form the neck of the mature phage, the channel for DNA injection. Since gp4 is the first protein to act after DNA packaging, the unstable DNA containing capsids from 4- -infected cells must represent the direct product of the packaging of DNA into procapsids. The major fraction of these capsids lost activity with a half-life of 1.1 minutes at 23 degrees C, though they were much more stable at 0 degree C. Electron microscopic observations indicated that the loss of activity was due to the DNA exiting from the incomplete capsids. The marginal stability of the condensed DNA molecules within capsids is consistent with models of ATP-driven condensation and spontaneous DNA ejection. The basis of the stability of these highly condensed molecules remains to be determined.
Links	PubMed
Keywords	Capsid/physiology; DNA, Viral/physiology; Microscopy, Electron; Models, Biological; Morphogenesis; Salmonella Phages/physiology; Salmonella typhimurium/physiology; Viral Envelope Proteins/physiology
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
BPP22:NEEDL		GO:0019073: viral DNA genome packaging	ECO:0000315:		P		Fro. I. Dependence of phage formation in vitro on the concentration of the capsid donor extract. Infected cell[ extracts containing incomplete capsids were freshly thawed and lysed as described in Materials and Methods. Diluted .samples were mixed with concentrated protein donor extract {5-) and incubated for I h at 23°C, at which time the reactions were stopped by further dilution and titered for viable phage. The background of the protein donor extract was 102 phage/mi. The background of the • at mo.~l, Conc-entrated calmid donor extracts were (0) 3.3× 10 (4- extract), (t) 2:5 × l0 T (10- extract) and (at) 2 x 103 (26-extract). Dependence of in vitro assembly on the concentration of the 4", lO ÷ and 26* activities. (a) Complementation of 10- extracts (O), 26- extracts (11) and 4- extracts (~lk) with protein donor extract. The capsid extracts were diluted 25-fold, to a final infected cell conventration of about 2 x 10 ° c~lls/ml. Samples were mixed with serial dilutions of the 5- protein donor. The maximum slo|~s determined from these curve,a were: 4+ activity, 5 to 6; 26 + activity. I to 2; 10 + activity, 4 to 5. The background of the undiluted extracts were protein donor 10a; 4- extract < 10s; 10- extracts, 5 x 10~; 26- extract, 2 x l0 a. (b) Variation of the ratio of cap`aid,a to protein; complemcntation of 4- capsids with cap,aid donor cell concentration: (O) 5x lO 9 cell,a/ml; (A) 2 x l0 s cell,a/ml; (m) 4 x 10 v (~lls/ml. The protein donor extract was from the same batch a.s used in (a).	complete CACAO 12034
BPP22:EXLYS		GO:0019073: viral DNA genome packaging	ECO:0000315:		P		Fro. I. Dependenc~ of phage formation in vitro on the concentration of the capsid donor extract. Infected ¢~zl[ extracts containing incomplete capsids were freshly thawed and lysed as described in Materials and Methods. D~uted .samples were mixed with concentrated protein donor extract {5-) and incubated for I h at 23°C, at which time the reactions were stopped by further dilution and titered for viable phage. The background of the protein donor extract was 102 phage/mi. The background of the • at mo.~l, Conc-entrated calmid donor extracts were (0) 3.3× 10 (4- extract), (t) 2:5 × l0 T (10- extract) and (at) 2 x 103 (26-extract). Fro. 4. Dependence of in vitro a.~.~em!~ly on the concentration of the 4", lO ÷ and 26* activities. (a) Com|)lementation of 10- extracts (O), 26- extracts (11) and 4- extracts (~lk) with protein donor extract. The cap,aid extracts were diluted 25-fold, to a final infected cell cont'entration of about 2 x 10 ° c~lls/ml. Samples were mixed with serial dilutions of the 5- prc)tein donor. The maximum slo|~s determined from these curve,a were: 4+ activity, 5 to 6; 26 + activity. I to 2; 10 + activity, 4 to 5. The background of the undiluted extracts were protein donor 10a; 4- extract < 10s; 10- extracts, 5 x 10~; 26- extract, 2 x l0 a. (b) Variation of the ratio of cap`aid,a to protein; complemcntation of 4- ~'apsids with cap,aid donor cell concentration: (O) 5x lO 9 cell,a/ml; (A) 2 x l0 s cell,a/ml; (m) 4 x i0 v (~lls/ml. The protein donor extract was from the same batch a.s used in (a).	complete CACAO 12032
BPP22:VG10		GO:0019073: viral DNA genome packaging	ECO:0000315:		P		Fro. 4. Dependence of in vitro a.~.~em!~ly on the concentration of the 4", lO ÷ and 26* activities. (a) Com|)lementation of 10- extracts (O), 26- extracts (11) and 4- extracts (~lk) with protein donor extract. The cap,aid extracts were diluted 25-fold, to a final infected cell cont'entration of about 2 x 10 ° c~lls/ml. Samples were mixed with serial dilutions of the 5- prc)tein donor. The maximum slo|~s determined from these curve,a were: 4+ activity, 5 to 6; 26 + activity. I to 2; 10 + activity, 4 to 5. The background of the undiluted extracts were protein donor 10a; 4- extract < 10s; 10- extracts, 5 x 10~; 26- extract, 2 x l0 a. (b) Variation of the ratio of cap`aid,a to protein; complemcntation of 4- ~'apsids with cap,aid donor cell concentration: (O) 5x lO 9 cell,a/ml; (A) 2 x l0 s cell,a/ml; (m) 4 x i0 v (~lls/ml. The protein donor extract was from the same batch a.s used in (a). ependenc~ of phage formation in vitro on the concentration of the capsid donor extract. Infected ¢~zl[ extracts containing incomplete capsids were freshly thawed and lysed as described in Materials and Methods. D~uted .samples were mixed with concentrated protein donor extract {5-) and incubated for I h at 23°C, at which time the reactions were stopped by further dilution and titered for viable phage. The background of the protein donor extract was 102 phage/mi. The background of the • at mo.~l, Conc-entrated calmid donor extracts were (0) 3.3× 10 (4- extract), (t) 2:5 × l0 T (10- extract) and (at) 2 x 103 (26-extract).	complete CACAO 12033

References

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