We describe the derivation of two strains of Salmonella typhimurium LT2 which are r-m+ for all three of the known chromosomal genes for the restriction and modification of DNA, hsdLT, hsdSA, and hsdSB; the strains were designated LB5000 and LB5010. LB5000 is a smooth derivative sensitive to phage P22; LB5010 is a galE strain sensitive to phage P1.

Stable restriction-deficient, modification-proficient galE (JRSO1) and F'galE+ (JR502) strains of Salmonella typhimurium were constructed and the effects of restriction on transformation by plasmid pBR322 were tested. Several factors which affect transformation efficiency were systematically examined to determine optimum transformation conditions and a simplified method is presented.

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Current genetic and molecular evidence places all the known type I restriction and modification systems of Escherichia coli and Salmonella enterica into one of four discrete families: type IA, IB, IC or ID. StySBLI is the founder member of the ID family. Similarities of coding sequences have identified restriction systems in E.coli and Klebsiella pneumoniae as probable members of the type ID family. We present complementation tests that confirm the allocation of EcoR9I and KpnAI to the ID family. An alignment of the amino acid sequences of the HsdS subunits of StySBLI and EcoR9I identify two variable regions, each predicted to be a target recognition domain (TRD). Consistent with two TRDs, StySBLI was shown to recognise a bipartite target sequence, but one in which the adenine residues that are the substrates for methylation are separated by only 6 bp. Implications of family relationships are discussed and evidence is presented that extends the family affiliations identified in enteric bacteria to a wide range of other genera.

Mutants ofEscherichia coli and Salmonella typhimurium that were deficient in protein methylesterase activity encoded by cheB had an inverted response to oxygen; they were repelled by concentrations of oxygen that attract wild-type bacteria. Normal responses to oxygen and phosphotransferase substrates were observed in mutants that were deficient in protein methyltransferase (CheR) and the methyl-accepting transducing proteins (Tsr, Tar, Trg). However, the methylation-independent response to oxygen was modified by the loss of esterase activity. The inversion was apparently effected by the amidated Tsr protein present in cheB tsr+ mutants because aerotaxis was normal in cheB tsr strains. Chemotaxis to phosphotransferase sugars was normal in cheB mutants provided the extreme clockwise bias of the flagellar motors was modified to increase the probability of counterclockwise rotation.Chemotaxis by Salmonella typhimurium and Escherichia coli toward most chemoeffectors involves methyl-accepting transducing proteins that span the cytoplasmic membrane (11,12,21,30,37). Attractant, or an attractant-receptor complex if the primary receptor is a soluble periplasmic protein, binds to a transducing protein at the outer surface, probably inducing a conformational change that is propagated to the interior of the cell and signals a decrease in the probability of clockwise rotation of the flagellar motors. The transducing protein is subsequently methylated by the cheR product, protein methyltransferase, and the net effect is to cancel the excitation signal so that the bacteria adapt to the new concentration of attractant (5, 31). If the attractant is withdrawn or a repellent is added to the adapted cells, the transducing protein sends a signal to the flagellar motor that greatly increases the probability of clockwise rotation. The transducing protein is then demethylated by the cheB product, protein methylesterase, until the signal for clockwise rotation is cancelled and the bacteria adapt (5, 33). The CheB protein is also involved in processing newly synthesized transducing proteins by glutaminyl residue deamidation which yields sites for methylesterification (8,25). Three known methyl-accepting transducing proteins are the products of the tsr, tar, and trg genes (6,10,27,29). The tap gene has extensive homology to tsr and tar, but no transducing role for the Tap protein has been demonstrated (12, 28).In addition to most chemoeffectors that act on methylaccepting transducing proteins, there are some attractants that are recognized by sensory transduction systems that are independent of methylation (19). Sugar substrates for the phosphotransferase transport system are attractants for one methylation-independent chemotactic system in which the enzyme II receptor for transport is also the receptor for chemotaxis (1). Adaptation to phosphotransferase sugars occurs normally in cells with impaired protein methylation * Corresponding author. t Present address: Snow-Brand Milk Products Co., Ltd., Research Institute of Life Science, 519, ...