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Lysozyme by devadatha


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lysozyme discovery and structure

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Lysozyme by devadatha

  1. 1. Submitted by; B.Devadatha M.Sc2nd sem Dept of Biochemistry and Molecular Biology. Submitted to; Dr. R. Rukkumani, Assistant Professor, Dept of Biochemistry and Molecular Biology. LYSOZYME
  2. 2.  Lysozyme: is 129 aminoacid residues enzyme  (EC, hydrolase which catalyzes hydrolysis of 1,4- beta-linkages between N-acetylmuramic acid and N-acetyl-D- glucosamine residues in peptidoglycan and between N-acetyl- D-glucosamine residues in chitodextrins.  Molecular weight of Lysozyme is an approximately 14.7 kDa. LYSOZYME
  3. 3. An enzyme found naturally in • egg white • human tears • saliva (as well as other body fluids) Capable of destroying the cell walls of certain bacteria. Acts as a mild antiseptic OCCURENCE
  4. 4. Historical background  Lysozyme was the first enzyme structure solved with X-ray crystallography (Structure 2LYZ in the PDB, deposited 1975 by Diamond and Phillips). It is now the most common enzyme in the PDB (966 structures), most of which are engineered mutants from the work of Matthews . Lashtchenko (1909) observed the antibacterial properties of hen egg white. Hen egg white lysozyme (HEWL) has become the “classic” lysozyme. Fleming (1922) noted the antibacterial property of nasal mucus.
  5. 5. FLEMINGS DISCOVERY OF LYSOZYME  The lysozyme was first noticed during some investigations made on a patient suffering from acute coryza  The nasal secretion of this patient was cultivated daily on blood agar plates and for the first three days of the infection there was no growth, with the exception of an occasional staphylococcus colony  The culture made from the nasal mucus on the fourth day showed in 24 hours a large number of small colonies which, on examination proved to be large gram-positive cocci .  The microbe has not been exactly identified alluded as  Micrococcus lysodeikticus.
  6. 6. PRELIMINARY EXPERIMENTS SHOWING THE ACTION OF THE LYSOZYME  Nasal mucus from the patient, with coryza, was shaken up with five times its volume of normal salt solution, and the mixture was centrifuged.  A drop of the clear supernatant fluid was placed on an agar plate, which had previously been thickly planted with M. lysodeikticus, and the plate was incubated at 37° C. for 24 hours.  It showed a copious growth of the coccus, except in the region where the nasal mucus has been placed.  There was complete inhibition of growth, and this inhibition extended for a distance of about 1 cm. Beyond the limits of the mucus.
  7. 7. The effect of the lysozyme on bacteria Inhibitory Action:  A small portion of the agar is removed from an ordinary agar plate making a cup into which some material rich in lysozyme  A drop of liquid agar, at a temperature of about 50° C., is placed on the material in the cup and is allowed to solidify after which the cup is filled with the liquid agar  Liquid agar is then poured all over the plate to make a thin layer over the original surface
  8. 8. • The whole surface of the medium is now thickly planted with the M. lysodeikticus and the plate is incubated for 24 hours. • when it will be seen that there is copious growth of the coccus except in the region of the implanted material. • It is evident that the lytic substance has dissolved the cocci for a distance of 3 or 4 mm.
  9. 9. Properties of lysozyme  Lysozyme has the characters of a ferment. The rapidity of its action increases up to 60° C, but at temperatures over 65° C. it is destroyed more or less rapidly.  It acts best in a neutral medium.  Peptic or tryptic digestion does not destroy lysozyme.  Stability—When kept dry, lysozyme can be preserved for a long time. It was noted that commercial dried egg albumen was very rich in lysozyme .
  10. 10. Distribution of lysozyme  In the human body:- Many tissues and secretions have been examined for lysozyme .  Of the secretions and body-fluids, all contained lysozyme except normal urine, sweat, and cerebrospinal fluid.  In the tissues of animals :- Florey, who titrated the lysozyme content of extracts of many tissues of several animals and compared them with human tissues.  Cat’s tissues, with the striking exception of the salivary gland, are relatively deficient in lysozyme.
  11. 11. CONCLUSION  lysozyme is a widely distributed antibacterial ferment which is probably inherent in all animal cells and constitutes a primary method of destroying bacteria  while acting most strikingly on non-pathogenic bacteria yet can, when allowed to act in the full strength in which it occurs in some parts of the body, attack pathogenic organisms.
  12. 12.  That it is very easy to make bacteria relatively resistant to lysozyme, so that any pathogenic microbe isolated from the body where it has been growing in the presence of a non-lethal concentration of lysozyme must have acquired increased resistance to the ferment.  There are some differences in the lysozyme of different tissues and in different animals whereby bacteria are susceptible to different lysozymes in varying degrees. CONCLUSION
  13. 13. The Hen Egg-white Lysozyme Molecule  The three-dimensional structure of the hen egg-white lysozyme molecule has been determined by X-ray analysis of the tetragonal crystals grown at pH 4.7 from 1 M sodium chloride solution.  The phases of the X-ray reflections were determined by the method of multiple isomorphous replacement developed to take systematic account of anomalous scattering effects .  It has been interpreted with reference to the known amino acid sequence of lysozyme as determined independently by Jolls and Canfield and their colleagues.  crystals of Lysozyme diffract X-Ray beam to a very high resolution, currently the highest resolution structure, presented in Protein Data Bank, was solved at resolution 0.94 Angstrom
  14. 14. Interpretation of the Electron Density Map • Hen egg-white lysozyme has a molecular weight of about 14,600 and each molecule comprises 129 amino acid residues. • The region of highest electron density, at lower right of center, was found to correspond to the half- cysteine residue 30. • since it clearly forms part of a helical conformation of the polypeptide chain, the axis of which is marked HH‘ • Four residues removed from an identifiable phenylalanine residue (34) nearer the carboxyl end of the chain
  15. 15. Role in disease • Lysozyme is part of the innate immune system. • Reduced lysozyme levels have been associated with broncho pulmonary dysplasia in newborns. • Since lysozyme is a natural form of protection from gram-positive pathogens like Bacillus and Streptococcus • In certain cancers (especially myelomonocytic leukemia) excessive production of lysozyme by cancer cells can lead to toxic levels of lysozyme in the blood. • High lysozyme blood levels can lead to kidney failure and low blood potassium.
  16. 16. Reference • On a Remarkable Bacteriolytic Element found in Tissues and Secretions. • By ALEXANDER FLEMING, M.B.,F.R.C.S. • THE HEN EGG-WHITE LYSOZYME MOLECULE • BY DAVID C. PHILLIPS
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