Identification and partial characterization of a novel hemolysin from Leptospira interrogans serovar lai

Seoung Hoon Lee, Kyung A. Kim, Yong Gun Park, In Wha Seong, Min Ja Kim, Yong Ju Lee

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Abstract

It has been suggested that leptospiral hemolysins are important in the virulence and pathogenesis of leptospirosis. We have isolated an Escherichia coli clone carrying the 7.8 kb DNA insert from a genomic library of Leptospira interrogans serovar lai by plaque hybridization using a sequence derived from the sphingomyelinase C gene (sphA) of L. borgpetersenii. The clone showed a clear β-hemolytic zone on sheep blood agar and high hemolytic activities on both human and sheep erythrocytes in liquid assays. The clone carried at least two genes responsible for the hemolytic activities, encoded by two open reading frames of 1662 and 816 nucleotides, which are named sphH and hap-1 (hemolysis associated protein-1), respectively. The SphH showed 75% homology to the SphA at the amino acid level, and the Hap-1 showed no significant homology in major databases. Interestingly, however, E. coli cells harboring sphH did not show sphingomyelinase or phospholipase activities. Moreover, SphH-mediated hemolysis was osmotically protected by polyethylene glycol 5000, suggesting that the hemolysis is likely to be caused by pore formation on the membrane. The SphH was successfully expressed in E. coli as a histidine (His)-SphH fusion protein. Both sphH and hap-1 were highly conserved among the Leptospira species, except for the absence of sphH in non-pathogenic L. biflexa serovar patoc. We concluded that the SphH is a novel hemolysin of a pathogenic Leptospira species, which may be a putative pore-forming protein. (C) 2000 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)19-28
Number of pages10
JournalGene
Volume254
Issue number1-2
DOIs
Publication statusPublished - 2000 Aug 22

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Keywords

  • Cloning
  • Nucleotide sequence
  • Pore-forming protein
  • hap-1
  • sphH

ASJC Scopus subject areas

  • Genetics

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