Regulation of Brucella abortus catalase as a defensive mechanism against oxidative stress

Abstract

Stinictiiral components of Brucella Pathogenesis of Brucella infection Oxidative stress in phagocytes Bacterial defensive mechanisms Catalase and superoxide dismutase Transcription factors of the LysR family 10 Rationale for this study References CHAPTER n. BRUCELLA ABORTUS PERIPLASMIC CATALASE IS REGULATED BY EXTERNAL HYDROGEN PEROXIDE 21 ABSTRACT INTRODUCTION 22 MATERIALS AND METHODS 24 Bacterial strains and medium 24 Construction of the deletion plasmid 24 Transformation of Brucella abortus 25 Halo assay (Hydrogen peroxide sensitivity assay) 25 2-D protein gel analysis 25 Western blot 26 Catalase assay 27 V Dot blot 27 RESUUrS Sensitivity of the catalase deletion mutant to HzCh 2-D gel analysis and identification of protein spots Response of Brucella abortus to oxidative stress 2-D gel analysis on Cu-Zn SOD deletion mutant 2-D gel analysis on catalase deletion mutant Enzyme activity Dot blot of catalase RNA DISCUSSION 31 REFERENCES CHAPTER in. OXYR REGULATES PERIPLASMIC CATALASE IN BRUCELLA ABORTUS

Identification of Brucella abortus OxyR and Its Role in Control of Catalase Expression

“…The procedure followed that given in Current Protocols in Molecular Biology (2) modified for Brucella (10). In this assay, a pronounced gel shift was observed in the presence of Brucella soluble extract (data not shown).…”

“…Brucella abortus has a single hydroperoxidase, a true catalase, which is expressed in the periplasm (17). In previous studies (10,11), it was shown that Brucella catalase is regulated at the transcriptional level.…”

“…In a preliminary study (10), primer extension indicated that transcription of the catalase gene begins at nucleotide 221 on the published catalase gene sequence (17). Immediately upstream are reasonable Ϫ10 and Ϫ35 promoter sequences, TAATTG and TGGAGA, respectively.…”