Genetics of Some Beneficial Traits ◾ 151
Drug-Delivery System
Many lactic acid bacteria are normally present in the gastrointestinal (GI) tract of humans and
food animals and birds, and some of them have beneficial effects on the health and well being of
the hosts (see Chapter 16). Currently, studies are in progress to anchor antigens of pathogens associated with enteric diseases in humans on the cell wall of suitable lactic acid bacteria (see Chapter 16).
The live cells then can be used as a drug-delivery vehicle to deliver antigens to stimulate antibody
production via the digestive tract. One method used to anchor proteins is to use Lac. lactis Prt P,
which normally remains attached to the cell wall with its carboxy-terminal domain. The effectiveness of the carboxy-terminal cell wall anchor of Staphylococcus aureus protein A to the cell wall of
Lac. lactis is also being investigated. Genes for certain virulence factors from enteric pathogens are
also cloned and expressed in lactic acid bacteria to provide protection against enteric diseases in
the gastrointestinal tract. 12
Production of Pediocin in Heterologous Hosts
The bacteriocin pediocin PA-1/AcH is produced by many strains of Ped. acidilactici. The molecule
is translated as a 62-amino acid prepediocin from which the 18-amino acid leader peptide at the
NH 2 terminal is enzymatically removed during membrane translocation through the ABC transporter system (Chapter 17). The 44-amino acid pediocin is then released in the growth medium.
The phenotype is plasmid linked, and the structural gene with three other genes is arranged in an
operon system, and the protein encoded by the last gene acts as the ABC transporter. The DNA
fragment with all four genes has been cloned in several Gram-negative vectors and in a shuttle
vector, and the host strains produce active pediocin molecules, generally at lower concentrations.
In a separate study, only the nucleotide segment of the matured pediocin (without the leader
segment) was fused to the carboxy terminus of the maltose-binding protein (MBP), a secretory
protein, and cloned in a Gram-negative vector. Escherichia coli transformed with the cloned vector
secreted the chimeric MBP–pediocin by the Sec-dependent transport system in the growth media.
The MBP–pediocin molecule, although much larger, retained the bactericidal property as that of
natural pediocin. The pediocin operon has also been cloned in suitable vectors and transformed in
Lac. lactis and Str. thermophilus strains, which produced pediocin. Generally, the production is low,
and the phenotype is unstable in the host in the absence of selective pressure, such as antibiotics (the
genes encoding the resistance to antibiotics are used as selective markers in the cloning vectors). 13
Protein Engineering
Many studies are being conducted to change the amino acid sequences of proteins of lactic acid
bacteria to determine the influence on the physical, chemical, and biological properties (structures
and functions) of the proteins.
Production of Hybrid Prepediocin
Like the prebacteriocin molecules of lactic acid bacteria, prepediocin molecules have an 18-amino
acid leader segment at the NH 2 terminus and a 44-amino acid segment at the carboxyl end.
Following translation, the leader peptide directs the molecule for translocation through the ABC
transporter. In one study, the promoter and nucleotide sequences of the leader segment of the
pediocin structural gene were replaced with a similar segment associated with the production of
Drug-Delivery System
Many lactic acid bacteria are normally present in the gastrointestinal (GI) tract of humans and
food animals and birds, and some of them have beneficial effects on the health and well being of
the hosts (see Chapter 16). Currently, studies are in progress to anchor antigens of pathogens associated with enteric diseases in humans on the cell wall of suitable lactic acid bacteria (see Chapter 16).
The live cells then can be used as a drug-delivery vehicle to deliver antigens to stimulate antibody
production via the digestive tract. One method used to anchor proteins is to use Lac. lactis Prt P,
which normally remains attached to the cell wall with its carboxy-terminal domain. The effectiveness of the carboxy-terminal cell wall anchor of Staphylococcus aureus protein A to the cell wall of
Lac. lactis is also being investigated. Genes for certain virulence factors from enteric pathogens are
also cloned and expressed in lactic acid bacteria to provide protection against enteric diseases in
the gastrointestinal tract. 12
Production of Pediocin in Heterologous Hosts
The bacteriocin pediocin PA-1/AcH is produced by many strains of Ped. acidilactici. The molecule
is translated as a 62-amino acid prepediocin from which the 18-amino acid leader peptide at the
NH 2 terminal is enzymatically removed during membrane translocation through the ABC transporter system (Chapter 17). The 44-amino acid pediocin is then released in the growth medium.
The phenotype is plasmid linked, and the structural gene with three other genes is arranged in an
operon system, and the protein encoded by the last gene acts as the ABC transporter. The DNA
fragment with all four genes has been cloned in several Gram-negative vectors and in a shuttle
vector, and the host strains produce active pediocin molecules, generally at lower concentrations.
In a separate study, only the nucleotide segment of the matured pediocin (without the leader
segment) was fused to the carboxy terminus of the maltose-binding protein (MBP), a secretory
protein, and cloned in a Gram-negative vector. Escherichia coli transformed with the cloned vector
secreted the chimeric MBP–pediocin by the Sec-dependent transport system in the growth media.
The MBP–pediocin molecule, although much larger, retained the bactericidal property as that of
natural pediocin. The pediocin operon has also been cloned in suitable vectors and transformed in
Lac. lactis and Str. thermophilus strains, which produced pediocin. Generally, the production is low,
and the phenotype is unstable in the host in the absence of selective pressure, such as antibiotics (the
genes encoding the resistance to antibiotics are used as selective markers in the cloning vectors). 13
Protein Engineering
Many studies are being conducted to change the amino acid sequences of proteins of lactic acid
bacteria to determine the influence on the physical, chemical, and biological properties (structures
and functions) of the proteins.
Production of Hybrid Prepediocin
Like the prebacteriocin molecules of lactic acid bacteria, prepediocin molecules have an 18-amino
acid leader segment at the NH 2 terminus and a 44-amino acid segment at the carboxyl end.
Following translation, the leader peptide directs the molecule for translocation through the ABC
transporter. In one study, the promoter and nucleotide sequences of the leader segment of the
pediocin structural gene were replaced with a similar segment associated with the production of