Cell lysis is responsible for the appearance of extracellular urease in Helicobacter pylori.
BACKGROUND
Helicobacter pylori is a neutralophilic bacterium that colonizes the acidic human gastric surface using the neutralizing capacity of a constitutively produced urease. Urease is present both in the cytoplasm and bound to the outside surface of the bacteria. The origin of the surface urease continues to be controversial. This study provides additional evidence that the origin of surface urease is cell lysis, not secretion.


METHODS
H. Pylori was transformed with a plasmid encoding green fluorescent protein (GFP), a non-native cytoplasmic protein. Cultures supplemented with beta-cyclodextrin or horse serum were collected over various time periods and spun through a ficoll cushion to gently separate whole bacteria from released protein. The pellet and supernatant fractions were analyzed by fluorimetry, SDS-PAGE and Coomassie blue or Western analysis.


RESULTS
GFP fluorescence and antigenic reactivity in the supernatant increased at each time point. GFP, the non-native cytoplasmic protein, and UreB, a native cytoplasmic protein, increased over time in the supernatant and both proteins were always present in the pellet fraction. UreI, an inner membrane protein, was only present in the pellet fraction. beta-galactosidase, a protein not found in H. pylori, was used as a negative control.


CONCLUSIONS
Since it is unlikely that there is an intrinsic secretion system for GFP, a non-native protein, its increasing presence over time in the supernate fraction along with UreB, and retention of UreI in the pellet fraction implies that cell lysis accounts for the presence of urease on the surface of H. pylori.
Marcus
E A
EA
Department of Physiology, University of California, Los Angeles and VA Greater Los Angeles Health Care System, Los Angeles, CA 90073, USA.
Scott
D R
DR
D001433:Bacteriolysis; D049452:Green Fluorescent Proteins; D016480:Helicobacter pylori / Q000201:enzymology*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D014510:Urease / Q000378:metabolism*
D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins; D014510:Urease