<?xml version="1.0" encoding="UTF-8"?><xml><records><record><source-app name="Biblio" version="7.x">Drupal-Biblio</source-app><ref-type>17</ref-type><contributors><authors><author><style face="normal" font="default" size="100%">Pandya, Vaibhav Kumar</style></author><author><style face="normal" font="default" size="100%">Shankar, S. Shiva</style></author><author><style face="normal" font="default" size="100%">Rajesh, S.</style></author><author><style face="normal" font="default" size="100%">Rathore, Rajeshwari</style></author><author><style face="normal" font="default" size="100%">Kumaran, Sangaralingam</style></author><author><style face="normal" font="default" size="100%">Kulkarni, Mahesh J.</style></author></authors></contributors><titles><title><style face="normal" font="default" size="100%">Mechanistic insights on anserine hydrolyzing activities of human carnosinases</style></title><secondary-title><style face="normal" font="default" size="100%">Biochemica ET Biophysica Acta- General Subjects </style></secondary-title></titles><keywords><keyword><style  face="normal" font="default" size="100%">Anserine</style></keyword><keyword><style  face="normal" font="default" size="100%">Carnosinase</style></keyword><keyword><style  face="normal" font="default" size="100%">Carnosine</style></keyword><keyword><style  face="normal" font="default" size="100%">Diabetes</style></keyword><keyword><style  face="normal" font="default" size="100%">LC-MS</style></keyword><keyword><style  face="normal" font="default" size="100%">MS</style></keyword><keyword><style  face="normal" font="default" size="100%">Multiple reaction monitoring (MRM)</style></keyword><keyword><style  face="normal" font="default" size="100%">Promiscuity</style></keyword></keywords><dates><year><style  face="normal" font="default" size="100%">2023</style></year><pub-dates><date><style  face="normal" font="default" size="100%">MAR</style></date></pub-dates></dates><volume><style face="normal" font="default" size="100%">1867</style></volume><language><style face="normal" font="default" size="100%">eng</style></language><abstract><style face="normal" font="default" size="100%">&lt;p&gt;
	Anserine and carnosine represent histidine-containing dipeptides that exert a pluripotent protective effect on human physiology. Anserine is known to protect against oxidative stress in diabetes and cardiovascular diseases. Human carnosinases (CN1 and CN2) are dipeptidases involved in the homeostasis of carnosine. In poikilothermic vertebrates, the anserinase enzyme is responsible for hydrolyzing anserine. However, there is no specific anserine hydrolyzing enzyme present in humans. In this study, we have systematically investigated the anserine hydrolyzing activity of human CN1 and CN2. A targeted multiple reaction monitoring (MRM) based approach was employed for studying the enzyme kinetics of CN1 and CN2 using carnosine and anserine as substrates. Surprisingly, both CN1 and CN2 can hydrolyze anserine effectively. The observed catalytic turnover rate (Vmax/[E]t) was 21.6 s-1 and 2.8 s-1 for CN1 and CN2, respectively. CN1 is almost eight-fold more efficient in hydrolyzing anserine compared to CN2, which is comparable to the efficiency of the carnosine hydrolyzing activity of CN2. The Michaelis constant (Km) value for CN1 (1.96 mM) is almost three-fold lower compared to CN2 (6.33 mM), representing higher substrate affinity for anserine-CN1 interactions. Molecular docking studies showed that anserine binds at the catalytic site of the carnosinases with an affinity similar to carnosine. Overall, the present study elucidated the inherent promiscuity of human carnosinases in hydrolyzing anserine using a sensitive LCMS/MS approach.&lt;/p&gt;
</style></abstract><issue><style face="normal" font="default" size="100%">3</style></issue><work-type><style face="normal" font="default" size="100%">Article</style></work-type><custom3><style face="normal" font="default" size="100%">&lt;p&gt;
	Foreign&lt;/p&gt;
</style></custom3><custom4><style face="normal" font="default" size="100%">&lt;p&gt;
	3&lt;/p&gt;
</style></custom4></record><record><source-app name="Biblio" version="7.x">Drupal-Biblio</source-app><ref-type>17</ref-type><contributors><authors><author><style face="normal" font="default" size="100%">Rajesh, S.</style></author><author><style face="normal" font="default" size="100%">Jathar, Swaraj</style></author><author><style face="normal" font="default" size="100%">Banarjee, Reema</style></author><author><style face="normal" font="default" size="100%">Sharma, Monika</style></author><author><style face="normal" font="default" size="100%">Palkar, Shivani</style></author><author><style face="normal" font="default" size="100%">Shankar, S. Shiva</style></author><author><style face="normal" font="default" size="100%">Kulkarni, Mahesh J.</style></author></authors></contributors><titles><title><style face="normal" font="default" size="100%">Simple freeze-thaw based method for efficient purification of recombinant human proinsulin from inclusion bodies</style></title><secondary-title><style face="normal" font="default" size="100%">Protein Expression and Purification</style></secondary-title></titles><keywords><keyword><style  face="normal" font="default" size="100%">Diabetes</style></keyword><keyword><style  face="normal" font="default" size="100%">Disulfide mapping</style></keyword><keyword><style  face="normal" font="default" size="100%">glucose</style></keyword><keyword><style  face="normal" font="default" size="100%">insulin</style></keyword><keyword><style  face="normal" font="default" size="100%">Mass spectrometry</style></keyword></keywords><dates><year><style  face="normal" font="default" size="100%">2025</style></year><pub-dates><date><style  face="normal" font="default" size="100%">MAR</style></date></pub-dates></dates><volume><style face="normal" font="default" size="100%">227</style></volume><pages><style face="normal" font="default" size="100%">106645</style></pages><language><style face="normal" font="default" size="100%">eng</style></language><abstract><style face="normal" font="default" size="100%">&lt;p&gt;
	Insulin is a pivotal peptide hormone essential for regulating glucose homeostasis. It has been known for over 100 years, but its production and purification methods are still under improvement. Escherichia coli based bacterial expression system is primarily used for insulin production. The human insulin protein expressed in bacteria usually forms inclusion bodies, complicating the purification process. Traditionally, insulin purification is a timeconsuming process involving urea-based denaturation methods, and various refolding techniques, followed by extensive chromatographic methods. Here, we report an easy and efficient purification of human proinsulin involving freeze-thaw based solubilization method. The extracted proinsulin inclusion bodies are treated with different concentrations of urea, followed by a freeze-thaw based solubilization. The freezing was carried out at various temperatures, mainly -80 degrees C, -20 degrees C, and -196 degrees C to determine the optimum condition for solubilization. Highest solubilization of proinsulin from the inclusion body was achieved with 0.5M urea and -20 degrees C. Further Nickel NTA-based purification was performed, and the purified protein was characterized for disulfide mapping by high-resolution mass spectrometer (HRMS). We also performed glucose uptake assays to validate the functional properties of purified proinsulin. This freeze-thaw based mild solubilization approach is a fast and effective method for getting bioactive proinsulin, which will help further design better purification and processing strategies for insulin production.&lt;/p&gt;
</style></abstract><work-type><style face="normal" font="default" size="100%">Article</style></work-type><custom3><style face="normal" font="default" size="100%">&lt;p&gt;Foreign&lt;/p&gt;
</style></custom3><custom4><style face="normal" font="default" size="100%">&lt;p&gt;
	1.4&lt;/p&gt;
</style></custom4></record></records></xml>