Related Experiment Videos
Genetic study of a membrane protein: DNA sequence alterations due to 17 lamB point mutations affecting adsorption of
J M Clément1, E Lepouce, C Marchal
1Unité de Programmation Moléculaire et Toxicologie Génétique, CNRS LA 271, INSERM U.163, Institut Pasteur, Paris, France.
Abstract:
Gene lamB encodes the outer membrane receptor for phage lambda in Escherichia coli K12. We have determined the DNA sequence alterations of 17 lamB point mutations which result in resistance to phage lambda h+. The mutations correspond to four phenotypic classes according to the pattern of growth of three phages which use the lambda receptor: lambda h (a one-step host-range derivative of lambda h+), lambda hh* (a two-step host-range derivative of lambda h+) and K10 (another lambdoid phage). Fourteen mutations are of the missense type and correspond to Gly to Asp changes distributed as follows. One class I mutation is at position 382 of the mature lambda receptor. Seven class I* mutations, four of which at least are independent, are at position 401. Six independent class II mutations are at position 151. The three other (class III) mutations are of the nonsense type. They change codons TGG (Trp) into TAG (amber) at positions 120 (two mutations) and 351 (one mutation). Implications of these results for the topological organization of the lambda receptor as well as possible reasons for the limited number of altered sites detected are discussed.
Insights
Researchers identified specific DNA changes in the lamB gene conferring resistance to phage lambda in Escherichia coli. These mutations affect the lambda receptor protein, impacting phage interactions and providing insights into receptor structure.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- The lamB gene in Escherichia coli K12 encodes the outer membrane receptor crucial for phage lambda entry.
- Bacterial outer membrane proteins play vital roles in cellular processes and interactions with external agents like bacteriophages.
Purpose of the Study:
- To determine the DNA sequence alterations in lamB conferring resistance to phage lambda.
- To correlate these mutations with phenotypic classes based on phage growth patterns.
- To understand the topological organization of the lambda receptor.
Main Methods:
- DNA sequencing of 17 lamB point mutations.
- Phenotypic analysis of mutant strains using three different phages (lambda h, lambda hh*, K10).
- Analysis of missense (Gly to Asp) and nonsense (Trp to amber) mutations.
Main Results:
- Identified 17 lamB mutations conferring phage lambda resistance.
- Four phenotypic classes were observed, linked to specific mutation sites.
- Missense mutations (Gly to Asp) occurred at positions 151, 382, and 401.
- Nonsense mutations (Trp to amber) occurred at positions 120 and 351.
Conclusions:
- Specific amino acid changes in the lambda receptor protein are responsible for phage resistance.
- The identified mutation sites provide insights into the structural and topological organization of the lambda receptor.
- The limited number of altered sites suggests specific functional regions within the receptor are critical for phage binding.