General method for fine mapping of the Escherichia coli K-12 lamB gene: localization of missense mutations affecting bacteriophage lambda adsorption

Abstract

lamB is the structural gene for the bacteriophage lambda receptor, a multifunctional protein located in the outer membrane of Escherichia coli K-12. We present a method for deletion mapping of any lamB mutations with a recognizable pheno-type. This method involves a transducing phage constructed by in vitro recombination which can also be used for complementation, deoxyribonucleic acid sequence, and in vitro protein synthesis studies with the mutated lamB gene. Using this method, we mapped 18 lamB missense mutations which confer resistance to phage lambda h+ (wild-type host range). The main results were the following. (i) None of the 18 mutations was located in the first 4 deletion intervals out of the 11 of the genetic map. (ii) These mutations were clustered according to their phenotype as follows. (a) Class I mutations, which allow growth of lambda h and lambda hh* (one-step and two-step host range mutants of lambda, respectively), were located in three regions--three in interval V, four in interval VIII-IX, and three in interval X-XI. Only the last three mutations still allowed growth of phage K10 which also uses the lambda receptor, and two of them still allowed reversible binding of lambda h+. (b) All seven class II mutations allowed only growth of lambda hh* and mapped in interval V. These results are discussed in the frame of a genetic approach to the functional topology of the lambda receptor.