Studies on Mycobacterial Response to Low Iron and Oxidative Stress by Proteomics

Tuberculosis (TB) is a devastating disease affecting approximately nine million people every year globally. Even though the disease progression for TB is known, the molecular mechanisms underlying the interactions between Mycobacterium tuberculosis and the host are not completely understood. Proteomics has been used for understanding these interactions between mycobacteria and the host. However, classical proteomic studies have concentrated only on investigating the differences in relative abundances of proteins between two different states of M. tuberculosis and dynamics of differential regulation have largely been ignored. In this study, mycobacterial stress response was studied from the point of protein turnover. Protein turnover is defined in this study as the ratio of synthesis over degradation. Studies were conducted to analyze the protein turnover profiles of differentially regulated proteins in mycobacteria exposed to low iron conditions. The results showed that many proteins showed decreased protein turnover between the low iron and high iron conditions. A comparative study of protein turnover and protein abundance was then carried out to analyze the response to M. tuberculosis exposed to low iron conditions. The comparison showed that protein turnover study can discern much more subtle changes than protein abundance studies alone. Application of Principal Component Analysis to protein abundance and turnover measurements showed that protein turnover measurements lie in a distinct dimension from abundance measurements. Hotelling’s T2 analysis was also carried out which combined both the abundance and turnover measurements to provide a gross index of change between the stressed and non-stressed conditions. Relative abundance measurements were then carried out to understand the nature of mycobacterial response to an integrated stress (IS) model comprising of both low iron and oxidative stress factors. Under the IS model, catalase peeroxidase (KatG) a primary antioxidant protein and superoxide dismutase (SOD) which are dependent on the availability of iron were found to be downregulated. Alkylhydroperoxide reductases (AhpC and AhpD) were found to be upregulated supporting the hypothesis that in the presence of low iron stress and oxidative stress, M. tuberculosis upregulates AhpC and AhpD when KatG and SOD are unavailable for negotiating with oxidative stresses.