Department of Biochemistry

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Browsing Department of Biochemistry by Author "Ahmed, Niyaz"
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    Iron-dependent RNA-binding activity of Mycobacterium tuberculosis aconitase
    ( 2007-06-01) Banerjee, Sharmistha ; Nandyala, Ashok Kumar ; Raviprasad, Podili ; Ahmed, Niyaz ; Hasnain, Seyed E.
    Cellular iron levels are closely monitored by iron regulatory and sensor proteins of Mycobacterium tuberculosis for survival inside macrophages. One such class of proteins systematically studied in eukaryotes and reported in a few prokaryotes are the iron-responsive proteins (IRPs). These IRPs bind to iron-responsive elements (IREs) present at untranslated regions (UTRs) of mRNAs and are responsible for posttranscriptional regulation of the expression of proteins involved in iron homeostasis. Amino acid sequence analysis of M. tuberculosis aconitase (Acn), a tricarboxylic acid (TCA) cycle enzyme, showed the presence of the conserved residues of the IRP class of proteins. We demonstrate that M. tuberculosis Acn is bifunctional. It is a monomeric protein that is enzymatically active in converting isocitrate to cis-aconitate at a broad pH range of 7 to 10 (optimum, pH 8). As evident from gel retardation assays, M. tuberculosis Acn also behaves like an IRP by binding to known mammalian IRE-like sequences and to predicted IRE-like sequences present at the 3′ UTR of thioredoxin (trxC) and the 5′ UTR of the iron-dependent repressor and activator (ideR) of M. tuberculosis. M. tuberculosis Acn when reactivated with Fe2+ functions as a TCA cycle enzyme, but upon iron depletion by a specific iron chelator, it behaves like an IRP, binding to the selected IREs in vitro. Since iron is required for the Acn activity and inhibits the RNA-binding activity of Acn, the two activities of M. tuberculosis Acn are mutually exclusive. Our results demonstrate the bifunctional nature of M. tuberculosis Acn, pointing to its likely role in iron homeostasis. Copyright © 2007, American Society for Microbiology. All Rights Reserved.
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    Molecular characterization of multidrug-resistant isolates of Mycobacterium tuberculosis from patients in North India
    ( 2002-02-02) Siddiqi, Noman ; Shamim, Mohammed ; Hussain, Seema ; Kumar Choudhary, Rakesh ; Ahmed, Niyaz ; Prachee, ; Banerjee, Sharmistha ; Savithri, G. R. ; Alam, Mahfooz ; Pathak, Niteen ; Amin, Amol ; Hanief, Mohammed ; Katoch, V. M. ; Sharma, S. K. ; Hasnain, Seyed E.
    The World Health Organization has identified India as a major hot-spot region for Mycobacterium tuberculosis infection. We have characterized the sequences of the loci associated with multidrug resistance in 126 clinical isolates of M. tuberculosis from India to identify the respective mutations. The loci selected were rpoB (rifampin), katG and the ribosomal binding site of inhA (isoniazid), gyrA and gyrB (ofloxacin), and rpsL and rrs (streptomycin). We found known as well as novel mutations at these loci. Few of the mutations at the rpoB locus could be correlated with the drug resistance levels exhibited by the M. tuberculosis isolates and occurred with frequencies different from those reported earlier. Missense mutations at codons 526 to 531 seemed to be crucial in conferring a high degree of resistance to rifampin. We identified a common Arg463Leu substitution in the katG locus and certain novel insertions and deletions. Mutations were also mapped in the ribosomal binding site of the inhA gene. A Ser95Thr substitution in the gyrA locus was the most common mutation observed in ofloxacin-resistant isolates. A few isolates showed other mutations in this locus. Seven streptomycin-resistant isolates had a silent mutation at the lysine residue at position 121. While certain mutations are widely present, pointing to the magnitude of the polymorphisms at these loci, others are not common, suggesting diversity in the multidrug-resistant M. tuberculosis strains prevalent in this region. Our results additionally have implications for the development of methods for multidrug resistance detection and are also relevant in the shaping of future clinical treatment regimens and drug design strategies.
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    Whole genome fingerprinting and genotyping of multiple drug resistant (MDR) isolates of Pseudomonas aeruginosa from endophthalmitis patients in India
    ( 2002-05-13) Ahmed, Niyaz ; Bal, Abhijit ; Khan, Aleem A. ; Alam, Mahfooz ; Kagal, Anju ; Arjunwadkar, Vidya ; Rajput, Amarsingh ; Majeed, Ahmed A. ; Rahman, Syed Asad ; Banerjee, Sharmistha ; Joshi, Suvarna ; Bharadwaj, Renu
    Genome sequence-based fluorescent amplified-fragment length polymorphism (FAFLP) analysis was investigated for fingerprinting and subtyping 42 multiple drug resistant (MDR) isolates of Pseudomonas aeruginosa from post-surgical endophthalmitis. The FAFLP profiles derived from EcoR1/Mse1 restricted fragments differentiated clinical isolates and were found to be extremely reproducible. Seventeen different amplification patterns (amplitypes) were observed for all the 42 isolates from 11 different patients. Also, we were able to genotype the isolates based on the differential amplification of a total of 31 FAFLP markers representing genomic fragments from the P. aeruginosa chromosome. This data appears to provide clues to the genetic diversity of endopthalmitis associated strains and could be converted into digitised fingerprints suitable for electronic manipulations and archiving. © 2002 Elsevier Science B.V. All rights reserved.