Studies in the development and evaluation of in-house molecular tools to identify and characterise the Mycobacterium tuberculosis in resource limited settings

Munish Puri developed and validated in-house molecular based tools to identify and characterise the Mycobacterium tuberculosis (Mtb) from rural PNG. He found that TaqMan based assays could be used to identify the Mtb and that high resolution melt may play a role in screening for drug resistance mutations. These outcomes could be further developed to provide cost effective approaches to lab-based diagnosis of TB in resource poor settings

Support vector machine (SVM) based multiclass prediction with basic statistical analysis of plasminogen activators

Plasminogen (Pg), the precursor of the proteolytic and fibrinolytic enzyme of blood, is converted to the active enzyme plasmin (Pm) by different plasminogen activators (tissue plasminogen activators and urokinase), including the bacterial activators streptokinase and staphylokinase, which activate Pg to Pm and thus are used clinically for thrombolysis. The identification of Pg-activators is therefore an important step in understanding their functional mechanism and derives new therapies

Characterization of Type 1 ribosome inactivating proteins in edible plants

The ribosome inactivating proteins (RIPs) from plants possess RNA N-glycosidase activity that depurinates the major rRNA, thus damaging ribosome in an irreversible manner and arresting protein synthesis. RIPs occur in fungi, bacteria and plants and are abundant in angiosperms, where they appear to have defensive role. RIPs are presently classified as rRNA N-glycosidase in the enzyme nomenclature (EC 3.2.2.22) and do exhibit other enzymatic activities such as ribonuclease and deoxyribonuclease activities. RIPs are classified into two groups based on their difference in their primary structure. Type I RIPs consist of a single polypeptide chain of approximately 26&ndash;35 kDa that possess an RNA N-glycosidase activity. These proteins have attracted a great deal of attention because of their anti-viral, anti-tumor, and anti-microbial activities, which is useful in medical research and development. Here, we describe isolation of a novel protein from Momordica sp, a highclimbing vine from family Cucurbitaceae which is native to the tropical regions of Africa, Asia, Arabia and Caribbean. The purified protein has been verified by SDS-PAGE and mass spectrometry to contain only single chain Type-1 ribosome inactivating proteins (RIPs). With present experiments, we determined the presence of RIPs in edible plant materials, including some that are eaten raw by human beings. The novel protein is further characterized to validate its therapeutic potential.<br /

Partial purification and characterization of exoinulinase from kluyveromyces marxianus YS-1 for preparation of high-fructose syrup

An extracellular exoinulinase( 2, 1-&nbsp;&szlig;- D fructan fructanohydrolase, EC 3.2.1.7), which catalyzes the hydrolysis of inulin into fructose and glucose, was purified 23.5-fold by ethanol precipitation, followed by Sephadex G-100 gel permeation from a cell-free extract of Kluyveromyces marxianus YS-1. The partially purified enzyme exhibited considerable activity between pH 5 to 6, with an optimum pH of 5.5, while it remained stable(100%) for 3 h at the optimum temperature of 50&ordm; c. Mn2+ and Ca2+ produced a 2A-fold and 1.2-fold enhancement in enzyme activity, whereas Hg2+ and Ag2+&nbsp; completely inhibited the inulinase. A preparation of the partially purified enzyme effectively hydrolyzed inulin, sucrose, and raffinose, yet no activity was found with starch, lactose, and maltose. The enzyme preparation was then successfully used to hydrolyze pure inulin and raw inulin from Asparagus racemosus for the preparation of a high-fructose syrup. In a batch system, the exoinulinase hydrolyzed 84.8% of the pure inulin and 86.7% of the raw Asparagus racemosus inulin, where fructose represented 43.6mg/ml and 41.3mg/ml, respectively.<br /

Partial purification and characterization of limonoate dehydrogenase from Rhodococcus fascians for the degradation of limonin

An extracellular limonoate dehydrogenase was purified 10-fold from a cell-free extract of Rhodococcus fascians by ammonium sulfate precipitation, dialysis, and ultrafiltration. This purified dehydrogenase catalyzed theconversion of limonoate to 17-dehydrolimonoate. The enzyme showed optimum activity at pH 8.0 and 40oC, with Km value of 0.9 &micro;M, and requires Zn ions and sulfhydryl groups for catalytic action. The enzyme activity was inhibited by Hg2+ and NaN3 ions. The degradation of limonin (66%) in Kinnow mandarin juice was successfully demonstrated with partiallypurified limonoate dehydrogenase. With scale-up preparation of limonoate dehydrogenase, a successful debittering operation of fruit juices appears feasible.<br /

Processing of citrus peel for the extraction of flavonoids for biotechnological applications

Flavonoids are extra nutritional constituents that naturally occur in small quantities in plants. They are a family of polyphenolic compounds that are widespread in nature and are consumed as part of the human diet in significant amounts. The diversity in their structure and bioactivity of flavonoids make these compounds an interesting candidate for biotechnology based research. Extraction of flavonoids from citrus employing the use of various techniques such as chemical and physical methods is attempted in this write up. The biotechnological potential of flavonoids is not currently exploited to its maximum since extraction procedures are in developing phase. The current knowledge on the sources of citrus flavonoids and their potential activities in alleviating human health is also discussed

Combination of Balsamin and Flavonoids Induce Apoptotic Effects in Liver and Breast Cancer Cells

Flavonoids such as naringenin, quercetin, and naringin are known to exhibit anticancer properties. In this study, we examined the effects of these flavonoids on cell viability and apoptotic pathways of cancer cells, either singly or in combination with the type 1 ribosome inactivating protein, Balsamin. Treatment with flavonoids (naringenin, quercetin, and naringin) plus Balsamin for 48 h reduced HepG2 and MCF-7 cell viability, increased the activation of caspase-3 and -8, and induced apoptosis through up-regulation of pro-apoptotic genes and down-regulation of anti-apoptotic genes. Out of the three flavonoids tested, the Balsamin-Naringenin and Balsamin-Quercetin combinations appeared to be most effective compared to the Balsamin-Naringin combination. Balsamin combined with flavonoids also activated endoplasmic reticulum (ER)-stress&ndash;mediated apoptosis in breast cancer (MCF-7) cells, which was not activated by Balsamin treatment alone. These experimental results showed that Balsamin combined with flavonoids can reduce HepG2 and MCF-7 cells viability and induce apoptosis, which could be considered as a promising therapeutic approach to sensitize cells to Balsamin treatment, thereby improving its efficacy in breast or liver cancer therapy

Repurposing mucosal delivery devices for live attenuated tuberculosis vaccines

Tuberculosis (TB) remains one of the most lethal infectious diseases globally. The only TB vaccine approved by the World Health Organization, Bacille Calmette-Guérin (BCG), protects children against severe and disseminated TB but provides limited protection against pulmonary TB in adults. Although several vaccine candidates have been developed to prevent TB and are undergoing preclinical and clinical testing, BCG remains the gold standard. Currently, BCG is administered as an intradermal injection, particularly in TB endemic countries. However, mounting evidence from experimental animal and human studies indicates that delivering BCG directly into the lungs provides enhanced immune responses and greater protection against TB. Inhalation therapy using handheld delivery devices is used for some diseases and allows the delivery of drugs or vaccines directly into the human respiratory tract. Whether this mode of delivery could also be applicable for live attenuated bacterial vaccines such as BCG or other TB vaccine candidates remains unknown. Here we discuss how two existing inhalation devices, the mucosal atomization device (MAD) syringe, used for influenza vaccines, and the Respimat® Soft Mist™ inhaler, used for chronic obstructive pulmonary disease (COPD) therapy, could be repurposed for mucosal delivery of live attenuated TB vaccines. We also outline the challenges and outstanding research questions that will require further investigations to ensure usefulness of respiratory delivery devices that are cost-effective and accessible to lower- and middle-income TB endemic countries

Harnessing the evolutionary information on oxygen binding proteins through Support Vector Machines based modules

© The Author(s) 2018 This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Abstract Objectives The arrival of free oxygen on the globe, aerobic life is becoming possible. However, it has become very clear that the oxygen binding proteins are widespread in the biosphere and are found in all groups of organisms, including prokaryotes, eukaryotes as well as in fungi, plants, and animals. The exponential growth and availability of fresh annotated protein sequences in the databases motivated us to develop an improved version of “Oxypred” for identifying oxygen-binding proteins. Results In this study, we have proposed a method for identifying oxy-proteins with two different sequence similarity cutoffs 50 and 90%. A different amino acid composition based Support Vector Machines models was developed, including the evolutionary profiles in the form position-specific scoring matrix (PSSM). The fivefold cross-validation techniques were applied to evaluate the prediction performance. Also, we compared with existing methods, which shows nearly 97% recognition, but, our newly developed models were able to recognize almost 99.99 and 100% in both oxy-50 and 90% similarity models respectively. Our result shows that our approaches are faster and achieve a better prediction performance over the existing methods. The web-server Oxypred2 was developed for an alternative method for identifying oxy-proteins with more additional modules including PSSM, available at http://bioinfo.imtech.res.in/servers/muthu/oxypred2/home.html

Immobilized enzyme technology for debittering citrus fruit juices

There has been increased interest in the use of immobilized enzymes in fruit juice industry for debittering of citrus fruit juices due to their high efficiency to remove bitter flavonoids. The structure of naringin, responsible for immediate bitterness, and of limonin, responsible for &quot;delayed bitterness&quot; has been discussed. This chapter also discusses various attempts that have been made to immobilize enzymes on an appropriate support so as to enable their use in debittering of citrus fruit juices. These include physicochemical and enzyme biotechnological approaches which makes the fruit juice more acceptable and cost effective to the consumer. Despite of high volume of production of citrus fruits and fruit juices, suitable processes to produce non-bitter citrus juice by immobilized enzymes technology has not yet commercialized globally.<br /