Submitted by;B.DevadathaM.Sc2nd semDept of Biochemistry and Molecular Biology.

Submitted to;Dr. R. Rukkumani,Assistant Professor,Dept of Biochemistry and Molecular Biology.

LYSOZYME

Lysozyme: is 129 aminoacid residues enzyme

(EC 3.2.1.17), 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

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

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.

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.

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.

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

• 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.

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 .

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.

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.

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

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

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

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.

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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