rinky Rajput - Academia.edu (original) (raw)

Papers by rinky Rajput

Biochemical Journal, 2020

Drug repurposing is an alternative avenue for identifying new drugs to treat tuberculosis (TB). D... more Drug repurposing is an alternative avenue for identifying new drugs to treat tuberculosis (TB). Despite the broad-range of anti-tubercular drugs, the emergence of multi-drug-resistant and extensively drug-resistant strains of Mycobacterium tuberculosis (Mtb) H37Rv, as well as the significant death toll globally, necessitates the development of new and effective drugs to treat TB. In this study, we have employed a drug repurposing approach to address this drug resistance problem by screening the drugbank database to identify novel inhibitors of the Mtb target enzyme, DNA gyrase. The compounds were screened against the ATPase domain of the gyrase B subunit (MtbGyrB47), and the docking results showed that echinacoside, doxorubicin, epirubicin, and idarubicin possess high binding affinities against MtbGyrB47. Comprehensive assessment using fluorescence spectroscopy, surface plasmon resonance spectroscopy (SPR), and circular dichroism (CD) titration studies revealed echinacoside as a pot...

Enzyme and Microbial Technology, 2012

An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromato... more An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45 kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60 • C, respectively. It was thiol activated with two-and eight-fold enhancement in presence of 10 mM DTT and βmercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants, detergents, and oxidizing agents where a nearly 2-to 3-fold enhancement was observed in presence of H 2 O 2 and NaHClO 3. It hydrolyzed broad range of complex substrates including feather keratin, haemoglobin, fibrin, casein,and α-keratin. Analysis of amidolytic activity revealed that it efficiently cleaved phenylalanine → leucine → alanine-p-nitroanilides. It also cleaved insulin B chain between Val 2-Asn 3 , Leu 6-Cys 7 and His 10-Leu 11 residues.

The international journal of biochemistry & cell biology, Jan 20, 2018

Alzheimer's disease is a severe brain illness that causes vast numbers of nerve cells in the ... more Alzheimer's disease is a severe brain illness that causes vast numbers of nerve cells in the brain to die, driven by the production and deposition of amyloid beta (Aβ) peptides. Intrinsically disordered proteins (IDPs) generally lack stable structures and are abundant in nature. Aβ peptide is a well-known IDP with a wide range of oligomeric forms. Dysfunctions in Aβ lead to oligomerization, formation of fibrils, and neurodegenerative disorders or other forms of dementia. In this study, we used replica exchange molecular dynamics (REMD) to elucidate the roles of different osmolytes, particularly urea and trimethylamine N-oxide (TMAO), to study shifts in IDP populations. REMD samples the conformational space efficiently and at physiologically relevant temperatures, compared to conventional molecular dynamics that sample at a constant temperature. Urea is known to minimize the aggregation process, while TMAO is beneficial for its stabilizing action. The two osmolytes displayed char...

Applied Microbiology and Biotechnology, 2013

Keratinases are well-recognized enzymes with the unique ability to attack highly cross-linked, re... more Keratinases are well-recognized enzymes with the unique ability to attack highly cross-linked, recalcitrant structural proteins such as keratin. Their potential in environmental clean-up of huge amount of feather waste has been well established since long. Today, they have gained importance in various other biotechnological and pharmaceutical applications. However, commercial availability of keratinases is still limited. Hence, to attract entrepreneurs, investors and enzyme industries it is utmost important to explicitly present the market potential of keratinases through detailed account of its application sectors. Here, the application areas have been divided into three parts: the first one is dealing with the area of exclusive applications, the second emphasizes protease dominated sectors where keratinases would prove better substitutes, and the third deals with upcoming newer areas which still await practical documentation. An account of benefits of keratinase usage, existing market size, and available commercial sources and products has also been presented.

Pro-sequences were swapped in cis between keratinases from Bacillus licheniformis (Ker BL) and Ba... more Pro-sequences were swapped in cis between keratinases from Bacillus licheniformis (Ker BL) and Bacillus pumilus (Ker BP) to construct Ker ProBP-BL and Ker ProBL-BP, respectively. Expression of these keratinases was carried out constitutively by E. coli HB101-pEZZ18 system. They were characterized with respect to their parent enzymes, Ker BL and Ker BP, respectively. Ker ProBP-BL became more thermostable with a t 1/2 of 45 min at 80 • C contrary to Ker BL which was not stable beyond 60 • C. Similarly, the activity of Ker ProBP-BL on keratin and casein substrate, i.e. K:C ratio increased to 1.2 in comparison to 0.1 for Ker BL. Hydrolysis of insulin B-chain revealed that the cleavage sites increased to six from four in case of Ker ProBP-BL in comparison to Ker BL. However, cleavage sites decreased from seven to four in case of Ker ProBL-BP in comparison to the parent keratinase, Ker BP. Likewise, Ker ProBL-BP revealed altered pH and temperature kinetics with optima at pH 10 and 60 • C in comparison to Ker BP which had optima at pH 9 and 70 • C. It also cleaved soluble substrates with better efficiency in comparison to Ker BP with K:C ratio of 1.6. Pro-sequence mediated conformational changes were also observed in trans and were almost similar to the features acquired by the chimeras constructed in cis by swapping the pro-sequence region.

Biochemical Journal, 2020

Drug repurposing is an alternative avenue for identifying new drugs to treat tuberculosis (TB). D... more Drug repurposing is an alternative avenue for identifying new drugs to treat tuberculosis (TB). Despite the broad-range of anti-tubercular drugs, the emergence of multi-drug-resistant and extensively drug-resistant strains of Mycobacterium tuberculosis (Mtb) H37Rv, as well as the significant death toll globally, necessitates the development of new and effective drugs to treat TB. In this study, we have employed a drug repurposing approach to address this drug resistance problem by screening the drugbank database to identify novel inhibitors of the Mtb target enzyme, DNA gyrase. The compounds were screened against the ATPase domain of the gyrase B subunit (MtbGyrB47), and the docking results showed that echinacoside, doxorubicin, epirubicin, and idarubicin possess high binding affinities against MtbGyrB47. Comprehensive assessment using fluorescence spectroscopy, surface plasmon resonance spectroscopy (SPR), and circular dichroism (CD) titration studies revealed echinacoside as a pot...

Enzyme and Microbial Technology, 2012

An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromato... more An extracellular keratinase from Bacillus pumilus KS12 was purified by DEAE ion exchange chromatography. It was a 45 kDa monomer as determined by SDS PAGE analysis. It was found to be an alkaline, serine protease with pH and temperature optima of 10 and 60 • C, respectively. It was thiol activated with two-and eight-fold enhancement in presence of 10 mM DTT and βmercaptoethanol, respectively. In addition, its activity was stimulated in the presence of various surfactants, detergents, and oxidizing agents where a nearly 2-to 3-fold enhancement was observed in presence of H 2 O 2 and NaHClO 3. It hydrolyzed broad range of complex substrates including feather keratin, haemoglobin, fibrin, casein,and α-keratin. Analysis of amidolytic activity revealed that it efficiently cleaved phenylalanine → leucine → alanine-p-nitroanilides. It also cleaved insulin B chain between Val 2-Asn 3 , Leu 6-Cys 7 and His 10-Leu 11 residues.

The international journal of biochemistry & cell biology, Jan 20, 2018

Alzheimer's disease is a severe brain illness that causes vast numbers of nerve cells in the ... more Alzheimer's disease is a severe brain illness that causes vast numbers of nerve cells in the brain to die, driven by the production and deposition of amyloid beta (Aβ) peptides. Intrinsically disordered proteins (IDPs) generally lack stable structures and are abundant in nature. Aβ peptide is a well-known IDP with a wide range of oligomeric forms. Dysfunctions in Aβ lead to oligomerization, formation of fibrils, and neurodegenerative disorders or other forms of dementia. In this study, we used replica exchange molecular dynamics (REMD) to elucidate the roles of different osmolytes, particularly urea and trimethylamine N-oxide (TMAO), to study shifts in IDP populations. REMD samples the conformational space efficiently and at physiologically relevant temperatures, compared to conventional molecular dynamics that sample at a constant temperature. Urea is known to minimize the aggregation process, while TMAO is beneficial for its stabilizing action. The two osmolytes displayed char...

Applied Microbiology and Biotechnology, 2013

Keratinases are well-recognized enzymes with the unique ability to attack highly cross-linked, re... more Keratinases are well-recognized enzymes with the unique ability to attack highly cross-linked, recalcitrant structural proteins such as keratin. Their potential in environmental clean-up of huge amount of feather waste has been well established since long. Today, they have gained importance in various other biotechnological and pharmaceutical applications. However, commercial availability of keratinases is still limited. Hence, to attract entrepreneurs, investors and enzyme industries it is utmost important to explicitly present the market potential of keratinases through detailed account of its application sectors. Here, the application areas have been divided into three parts: the first one is dealing with the area of exclusive applications, the second emphasizes protease dominated sectors where keratinases would prove better substitutes, and the third deals with upcoming newer areas which still await practical documentation. An account of benefits of keratinase usage, existing market size, and available commercial sources and products has also been presented.

Pro-sequences were swapped in cis between keratinases from Bacillus licheniformis (Ker BL) and Ba... more Pro-sequences were swapped in cis between keratinases from Bacillus licheniformis (Ker BL) and Bacillus pumilus (Ker BP) to construct Ker ProBP-BL and Ker ProBL-BP, respectively. Expression of these keratinases was carried out constitutively by E. coli HB101-pEZZ18 system. They were characterized with respect to their parent enzymes, Ker BL and Ker BP, respectively. Ker ProBP-BL became more thermostable with a t 1/2 of 45 min at 80 • C contrary to Ker BL which was not stable beyond 60 • C. Similarly, the activity of Ker ProBP-BL on keratin and casein substrate, i.e. K:C ratio increased to 1.2 in comparison to 0.1 for Ker BL. Hydrolysis of insulin B-chain revealed that the cleavage sites increased to six from four in case of Ker ProBP-BL in comparison to Ker BL. However, cleavage sites decreased from seven to four in case of Ker ProBL-BP in comparison to the parent keratinase, Ker BP. Likewise, Ker ProBL-BP revealed altered pH and temperature kinetics with optima at pH 10 and 60 • C in comparison to Ker BP which had optima at pH 9 and 70 • C. It also cleaved soluble substrates with better efficiency in comparison to Ker BP with K:C ratio of 1.6. Pro-sequence mediated conformational changes were also observed in trans and were almost similar to the features acquired by the chimeras constructed in cis by swapping the pro-sequence region.