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Klebsiella pneumoniae is an important opportunistic gram-
negative pathogen that usually causes serious disease in
infants, the elderly and the immunocompromised host. It accounts
for a substantial percentage of nosocomal infections including
pneumonia, bacteremia, supprative infections, and urinary tract
and other infections in human. Although the exact mechanisms for
the pathogenesis of K. pneumonia are not fully understood,
several potential virulence factors have been described. One of
the potential virulence factors is hemolysin. It has been
suggested that hemolysin molecules may lyse other cells such as
leukocytes and macrophage resulting in lowered host resistance.
Rudenv, IA proposed hemolysin could be the major pathogenic
factor of K. pneumoniae in 1993. To study whether hemolysin
might implicate in pathogenic processes of K. pneumoniae
infections or not, our laboratory have cloned hemolysin gene
from K. pneumoniae. To further analyze the genetic information
of this gene, we processed a series of subcloning and finished
its restriction map, and then used sanger's dideoxy chain
termination method to determinate its nucleotide sequence.The
finished nucleotide sequence predicted two possible open reading
frames, the first one named as Kly A, and the later termed Kly
B. The nucleotide sequence and deduced amino acid sequence of
the Kly B gene showed a high homology to the entire region of
Legiolysin of Legionella pneumoniae. By the Construction of
internal deletion sabclones, we suggested that Kly B and
Legiolysin had the same propriety, they both could lyse
retinocyte and produce pigment-like material. Although the
deduced amino acid sequence of Kly A did not show any
significant homology to those of any other hemolysin ,we
suggested that the following experiment of Kly A should focus on
its hydrophobicity and homology to some channel like proteins.
In the other hand,it seems that there are some interactions such
as activation and modification between Kly A and Kly B. Becauses
of not only the distance of the genetic location of KlyA and
KlyB just 90 base pairs but also their phenotype on blood agar
plate is diffuse b-hemolytic zone when they are adjacent to each
other. Furthermore,with PET expression system，we had
overexpressed Kly B without N-terminal region and partial puri-
fication of that by High-Q column to prepare polycolonyl
antibody.Thought we got the protein wich was above 80% purity
and could be used to prepare polyclonyl antibody,finallly,the
results of western blotting showed that the preparation of
polyclonyl antibody was failed.
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