ABSTRACT
Changes in the structure of the Escherichia coli nucleoid during heat damage and repair were followed by sedimentation analysis in neutral sucrose gradients. Heating at 50 degrees C results first in a decrease in the sedimentation coefficient of the isolated nucleoid. Increasing the heating time, a subsequent increase in sedimentation coefficient is observed. After a heat shock (i.e. 4 min at 50 degrees C), a short incubation at 25 degrees C (i.e. 5 min) allows the nucleoid to repair and return to the sedimentation coefficient of control unheated nucleoids. The nucleoids heated at 50 degrees C for longer periods and incubated afterwards at 25 degrees C demonstrate a different pattern of structural repair. They associate with protein in the first stage of the repair period.
Subject(s)
Escherichia coli/ultrastructure , Hot Temperature , Bacterial Proteins/physiology , Centrifugation, Density Gradient , Chromosomes, Bacterial/physiology , DNA Repair , DNA, Bacterial/physiology , DNA, Superhelical/physiology , Escherichia coli/physiology , Kinetics , Organoids/physiologyABSTRACT
The repair of the Escherichia coli nucleoid structure after heat shock (50 degrees C, 5 min) was studied. After heat shock the repair process did not include the association of the nucleoid to protein structures as is the case after more severe heat treatments resulting in cell death or inactivation.
Subject(s)
Chromosomes, Bacterial/physiology , Escherichia coli/ultrastructure , Hot Temperature , Centrifugation, Density Gradient , Escherichia coli/physiologyABSTRACT
The effects of the centrifugal field on the sedimentation coefficient of the heated (50 degrees C, 30 min) Escherichia coli nucleoid were investigated. Form 3,000 r.p.m. the sedimentation coefficients of the heated nucleoids were highly dependent on rotor speed. At 3,000 r.p.m. thier sedimentation coefficient was about 4,000 S while at 7,000 r.p.m. if was about 1,500-1,700 S. At 7,000 r.p.m. and over, nucleoid aggregations occurred and it was difficult to differentiate speed dependence from nucleoid aggregation. Factors likely to cause speed dependence and/or nucleoid aggregation are indicated. The practical importance of these findings is pointed out.
Subject(s)
Chromosomes, Bacterial/analysis , Escherichia coli/ultrastructure , Bacterial Proteins , Centrifugation, Density Gradient , DNA, Bacterial , Hot Temperature , Nucleic Acid Conformation , Protein ConformationABSTRACT
The folded chromosome or nucleoid of Escherichia coli was analyzed by low-speed sedimentation in neutral sucrose gradients after heat treatment (30 min at 50 degrees C) and subsequent incubation of cells at 37 degrees C for various times. Heat treatment resulted in in vivo association of the nucleoids with cellular protein and in an increase in sedimentation coefficient. During incubation at 37 degrees C, a fraction of the nucleoids, from heated cells, because dissociated from cellular protein and regained their characteristic sedimentation coefficients. The percentage of nucleoids which returned to their control sedimentation position in the sucrose gradients corresponded to the percentage of cells able to repair thermal damage as assayed by enumeration on agar plates.
Subject(s)
Chromosomes, Bacterial/physiology , Escherichia coli/ultrastructure , Bacterial Proteins/analysis , Centrifugation, Density Gradient , Chromosomes, Bacterial/analysis , Escherichia coli/physiology , Hot TemperatureABSTRACT
The folded chromosome or nucleoid of Escherichia coli was analyzed by low-speed sedimentation in neutral sucrose gradients after in vivo heat treatment. Heat treatment of cultures at 50 degree C for 15, 30, and 60 min resulted in in vivo association of the nucleoids with cellular protein. Structural changes, determined by the increase in speed dependence of the nucleoids from heated cells, also occurred. These changes were most likely due to the unfolding of the typical compact nucleoid structure. The nucleoids from heated cells also had notably higher sedimentation coefficients (3,000 to 4,500S) than nucleoids from control cells (1,800S). These nucleoids did not contain greater than normal amounts of membrane phospholipids or ribonucleic acid. We propose that the protein associated with the nucleoids from heated cells causes the observed sedimentation coefficient increases.
