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B orrelia burgdorferi HtrA : evidence for twofold proteolysis of outer membrane protein p66
Author(s) -
Coleman James L.,
Toledo Alvaro,
Benach Jorge L.
Publication year - 2016
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/mmi.13221
Subject(s) - biology , periplasmic space , borrelia burgdorferi , lipid raft , bacterial outer membrane , membrane protein , microbiology and biotechnology , proteolysis , serine protease , protease , biochemistry , membrane , escherichia coli , enzyme , antibody , gene , immunology
Summary In prokaryotes, members of the H igh T emperature R equirement A ( HtrA ) family of serine proteases function in the periplasm to degrade damaged or improperly folded membrane proteins. B orrelia burgdorferi , the agent of Lyme disease, codes for a single HtrA homolog. Two‐dimensional electrophoresis analysis of B . burgdorferi B 31 A 3 and a strain that overexpresses HtrA ( A3HtrAOE ) identified a downregulated protein in A3HtrAOE with a mass, pI and MALDI‐TOF spectrum consistent with outer membrane protein p66. P 66 and HtrA from cellular lysates partitioned into detergent‐resistant membranes, which contain cholesterol‐glycolipid‐rich membrane regions known as lipid rafts, suggesting that HtrA and p66 may reside together in lipid rafts also. This agrees with previous work from our laboratory, which showed that HtrA and p66 are constituents of B . burgdorferi outer membrane vesicles. HtrA degraded p66 in vitro and A3HtrAOE expressed reduced levels of p66 in vivo . Fluorescence confocal microscopy revealed that HtrA and p66 colocalize in the membrane. The association of HtrA and p66 establishes that they could interact efficiently and their protease/substrate relationship provides functional relevance to this interaction. A3HtrAOE also showed reduced levels of p66 transcript in comparison with wild‐type B 31 A 3, indicating that HtrA ‐mediated regulation of p66 may occur at multiple levels.