application of alternative food-preservation - Bentham Science

Enzymes as Antimicrobials Application of Alternative Food-Preservation Technologies 59
Table 1: Different types of lysozyme.
Lysozyme types Provenience References
Conventional or chicken-type
c-lysozyme
-egg white of domestic chicken (Gallus gallus);
-purified from various tissues and secretions of mammals including
milk, saliva, tears, urine, respiratory and cervical secretions
Other types of lysozyme:
[3, 5, 9]
g-type lysozyme egg white of domestic goose [4, 9-11]
h-type lysozyme Plant
i-type lysozyme Invertebrates
b-type lysozyme Bacteria (Bacillus)
v-type lysozyme Viruses
Despite the variability in the amino acid composition and sequence of lysozyme molecules, amino acids of the
catalytic centre of the active site are well conserved [9].
In 1965 the structure of lysozyme was solved by X-Ray analysis with 2 angstrom resolution by David Chilton
Phillips. In particular, lysozyme structure is characterized by an α-helix which links two domains of the
molecule; one is mainly β-sheet in structure, and the other primarly α-helical. The hydrophilic groups are mainly
oriented inward, with most of the hydrophilic residues on the exterior of the molecule [12].
It has been proposed that the enzymatic action of the molecule is dependent on its ability to change the relative
position of its two domains and cause large conformational changes in the molecule.
In particular, glutamic acid and aspartic acid residues are directly involved in the breakdown of the glycosidic
bond between N-acetylglucosamine and N-acetylmuramic and their presence in the catalytic centre is thus
crucial for the hydrolytic activity of the enzyme. However, the amino acid sequence of known lysozymes reveals
that aspartic acid is not consistently present in the active site of lysozyme molecules [8]. In contrast, the
substitution of glutamic acid results in a complete inactivation of the enzyme [8], thus confirming the critical
role of this amino acid in the enzymatic activity of lysozyme [9].
ANTIMICROBIAL EFFECT AND MODE OF ACTION OF LYSOZYME
Lysozyme has been recognized to possess many physiological and functional properties, including important
roles in surveillance of membranes of mammalian cells; it enhances phagocytic activity of polymorphonuclear
leukocytes and macrophages and stimulates proliferation and antitumor functions of monocytes [7], but its high
microbicidal activity remains, by far, the main virtue that explains the high attention of scientists and industrial
stakeholders for its practical applications in medicine and food industry [9].
The antimicrobial activity of lysozyme has been extensively demonstrated in vitro or in physiological fluids and
secretions including milk, blood serum, saliva, and urine [13]. Although lysozyme has been shown to have
antimicrobial activities towards bacteria, fungi, protozoan and viruses [14-16], it is essentially known for its
antibacterial activity and has been used, on this basis, in food preservation.
Lysozyme has been demonstrated to be active throughout a wide pH range of 4–10; however, high ionic strength
(>0.2 M salt) was shown to have an inhibitory effect on lysozyme activity [2]. Under physiological conditions,
only a minority of Gram positive bacteria are susceptible to lysozyme, and it has been suggested that the main
role of lysozyme is to participate in the removal of bacterial cell walls, after the bacteria have been killed by
antimicrobial polypeptides present in egg albumin, insect hemolymph [17] or by complement in animal serum
[18]. This is in line with the notion that the lytic action of lysozyme does not kill susceptible bacteria under
physiological conditions, osmotically balanced [19].
The bacteriostatic and bactericidal properties of lysozyme have been the subject of many studies, and over the
last 10 years, several authors have proposed a novel antibacterial mechanism of action of lysozyme that is
independent of its muramidase activity [20, 21]. A non-lytic bactericidal mechanism involving membrane