Membrane-Bound Lytic Endotransglycosylase inEscherichia coli

ABSTRACT The gene for a novel endotype membrane-bound lytic transglycosylase, emtA, was mapped at 26.7 min of theE. coli chromosome. EmtA is a lipoprotein with an apparent molecular mass of 22 kDa. Overexpression of the emtA gene did not result in bacteriolysis in vivo, but the enzyme was shown to hydrolyze glycan strands isolated from murein by amidase treatment. The formation of tetra- and hexasaccharides, but no disaccharides, reflects the endospecificity of the enzyme. The products are characterized by the presence of 1,6-anhydromuramic acid, indicating a lytic transglycosylase reaction mechanism. EmtA may function as a formatting enzyme that trims the nascent murein strands produced by the murein synthesis machinery into proper sizes, or it may be involved in the formation of tightly controlled minor holes in the murein sacculus to facilitate the export of bulky compounds across the murein barrier.

[1]  J. Höltje,et al.  Purification and properties of a membrane-bound lytic transglycosylase from Escherichia coli , 1994, Journal of bacteriology.

[2]  J. Weiner,et al.  Complementation of growth defect in an ampC deletion mutant of Escherichia coli. , 1993, FEMS microbiology letters.

[3]  K. Isono,et al.  The physical map of the whole E. coli chromosome: Application of a new strategy for rapid analysis and sorting of a large genomic library , 1987, Cell.

[4]  S. Henikoff,et al.  Automated construction and graphical presentation of protein blocks from unaligned sequences. , 1995, Gene.

[5]  K. Mullis,et al.  Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. , 1988, Science.

[6]  W Keck,et al.  Peptidoglycan as a barrier to transenvelope transport , 1996, Journal of bacteriology.

[7]  H. Nojima,et al.  High efficiency transformation of Escherichia coli with plasmids. , 1990, Gene.

[8]  S. Mizushima,et al.  Accumulation of glyceride-containing precursor of the outer membrane lipoprotein in the cytoplasmic membrane of Escherichia coli treated with globomycin. , 1980, The Journal of biological chemistry.

[9]  E. Koonin,et al.  A family of lysozyme-like virulence factors in bacterial pathogens of plants and animals. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[10]  B. Walderich,et al.  Subcellular distribution of the soluble lytic transglycosylase in Escherichia coli , 1991, Journal of bacteriology.

[11]  B. Henrissat,et al.  Structures and mechanisms of glycosyl hydrolases. , 1995, Structure.

[12]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[13]  J. Yon-Kahn.  Lysozymes, model enzymes in biochemistry and biology , 1996 .

[14]  V. Braun,et al.  Chapter 14 Lipoproteins, structure, function, biosynthesis and model for protein export , 1994 .

[15]  K. H. Kalk,et al.  Structure of the 70-kDa soluble lytic transglycosylase complexed with bulgecin A. Implications for the enzymatic mechanism. , 1995, Biochemistry.

[16]  N. Sharon.  The bacterial cell wall. , 1969, Scientific American.

[17]  J. Höltje.  A hypothetical holoenzyme involved in the replication of the murein sacculus of Escherichia coli. , 1996, Microbiology.

[18]  W. Weidel,et al.  BAGSHAPED MACROMOLECULES--A NEW OUTLOOK ON BACTERIAL CELL WALLS. , 1964, Advances in enzymology and related subjects of biochemistry.

[19]  W. Keck,et al.  Identification of new members of the lytic transglycosylase family in Haemophilus influenzae and Escherichia coli. , 1996, Microbial drug resistance.

[20]  B. Lugtenberg,et al.  Electrophoretic resolution of the ‘major outer membrane protein’ of Escherichia coli K12 into four bands , 1975, FEBS letters.

[21]  M. de Pedro,et al.  Exoenzymatic activity of transglycosylase isolated from Escherichia coli. , 1981, European journal of biochemistry.

[22]  T. Beveridge,et al.  Analysis of the sodium dodecyl sulfate-stable peptidoglycan autolysins of select gram-negative pathogens by using renaturing polyacrylamide gel electrophoresis , 1994, Journal of bacteriology.

[23]  B. Glauner,et al.  Growth pattern of the murein sacculus of Escherichia coli. , 1990, The Journal of biological chemistry.

[24]  B. Glauner.  Separation and quantification of muropeptides with high-performance liquid chromatography. , 1988, Analytical biochemistry.

[25]  Thomas L. Madden,et al.  Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. , 1997, Nucleic acids research.

[26]  J. Höltje,et al.  Isolation and separation of the glycan strands from murein of Escherichia coli by reversed-phase high-performance liquid chromatography. , 1990, Analytical biochemistry.

[27]  T. Romeis,et al.  Specific interaction of penicillin-binding proteins 3 and 7/8 with soluble lytic transglycosylase in Escherichia coli. , 1994, The Journal of biological chemistry.

[28]  G. Shockman.  Microbial peptidoglycan (murein) hydrolases , 1994 .

[29]  A. Rodríguez-Tébar,et al.  Specific location of penicillin-binding proteins within the cell envelope of Escherichia coli , 1986, Journal of bacteriology.

[30]  B. Spratt.  Properties of the penicillin-binding proteins of Escherichia coli K12,. , 1977, European journal of biochemistry.

[31]  J. Mattick,et al.  'Touchdown' PCR to circumvent spurious priming during gene amplification. , 1991, Nucleic acids research.

[32]  U. Schwarz,et al.  Novel type of murein transglycosylase in Escherichia coli , 1975, Journal of bacteriology.

[33]  S. Cohen,et al.  Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. , 1980, Journal of molecular biology.

[34]  J. Höltje,et al.  Growth of the Stress-Bearing and Shape-Maintaining Murein Sacculus of Escherichia coli , 1998, Microbiology and Molecular Biology Reviews.

[35]  U. Schwarz,et al.  Escherichia coli murein transglycosylase. Purification by affinity chromatography and interaction with polynucleotides. , 1980, European journal of biochemistry.

[36]  J. Heijenoort,et al.  Bacterial‐Cell‐Wall Peptidoglycan Fragments Produced by Phage λ or Vi II Endolysin and Containing 1,6‐Anhydro‐N‐acetylmuramic Acid , 1975 .

[37]  B. Henrissat,et al.  Domains in microbial beta-1, 4-glycanases: sequence conservation, function, and enzyme families. , 1991, Microbiological reviews.

[38]  W. Vollmer,et al.  Characterization of three different lytic transglycosylases in Escherichia coli. , 1993, FEMS microbiology letters.

[39]  N. W. Davis,et al.  The complete genome sequence of Escherichia coli K-12. , 1997, Science.

[40]  A. Forsgren,et al.  Protein D, the immunoglobulin D-binding protein of Haemophilus influenzae, is a lipoprotein , 1992, Infection and immunity.

[41]  M. Templin,et al.  Cloning and expression of a murein hydrolase lipoprotein from Escherichia coli , 1995, Molecular microbiology.

[42]  Jeffrey H. Miller.  Experiments in molecular genetics , 1972 .

[43]  R. Eferl,et al.  Gene 19 of plasmid R1 is required for both efficient conjugative DNA transfer and bacteriophage R17 infection , 1995, Journal of bacteriology.

[44]  H. Rogers,et al.  Microbial cell walls and membranes , 1980 .

[45]  W. Vollmer,et al.  Outer membrane localization of murein hydrolases: MltA, a third lipoprotein lytic transglycosylase in Escherichia coli , 1997, Journal of bacteriology.

[46]  M. Inouye,et al.  A single amino acid determinant of the membrane localization of lipoproteins in E. coli , 1988, Cell.

[47]  U. Schwarz,et al.  Autolytic enzymes and cell division of Escherichia coli. , 1969, Journal of molecular biology.