Ravi Kanth Kamlekar
Bio: Ravi Kanth Kamlekar is an academic researcher from VIT University. The author has contributed to research in topics: Glycolipid transfer protein & Chemistry. The author has an hindex of 13, co-authored 25 publications receiving 490 citations. Previous affiliations of Ravi Kanth Kamlekar include University of Hyderabad & University of Minnesota.
TL;DR: A ubiquitously expressed lipid transfer protein, human GLTPD1, named here CPTP, is shown to specifically transfer C1P between membranes, triggering pro-inflammatory eicosanoid generation.
Abstract: Phosphorylated sphingolipids ceramide-1-phosphate (C1P) and sphingosine-1-phosphate (S1P) have emerged as key regulators of cell growth, survival, migration and inflammation. C1P produced by ceramide kinase is an activator of group IVA cytosolic phospholipase A2α (cPLA2α), the rate-limiting releaser of arachidonic acid used for pro-inflammatory eicosanoid production, which contributes to disease pathogenesis in asthma or airway hyper-responsiveness, cancer, atherosclerosis and thrombosis. To modulate eicosanoid action and avoid the damaging effects of chronic inflammation, cells require efficient targeting, trafficking and presentation of C1P to specific cellular sites. Vesicular trafficking is likely but non-vesicular mechanisms for C1P sensing, transfer and presentation remain unexplored. Moreover, the molecular basis for selective recognition and binding among signalling lipids with phosphate headgroups, namely C1P, phosphatidic acid or their lyso-derivatives, remains unclear. Here, a ubiquitously expressed lipid transfer protein, human GLTPD1, named here CPTP, is shown to specifically transfer C1P between membranes. Crystal structures establish C1P binding through a novel surface-localized, phosphate headgroup recognition centre connected to an interior hydrophobic pocket that adaptively expands to ensheath differing-length lipid chains using a cleft-like gating mechanism. The two-layer, α-helically-dominated 'sandwich' topology identifies CPTP as the prototype for a new glycolipid transfer protein fold subfamily. CPTP resides in the cell cytosol but associates with the trans-Golgi network, nucleus and plasma membrane. RNA interference-induced CPTP depletion elevates C1P steady-state levels and alters Golgi cisternae stack morphology. The resulting C1P decrease in plasma membranes and increase in the Golgi complex stimulates cPLA2α release of arachidonic acid, triggering pro-inflammatory eicosanoid generation.
TL;DR: Glycolipid transfer proteins originally were identified as small, soluble, amphitropic proteins that specifically accelerate the intermembrane transfer of glycolipids but now are known to adopt a unique, helically dominated, two-layer ‘sandwich’ architecture defined as the GLTP-fold that provides the structural underpinning for the eukaryotic GLTP superfamily.
Abstract: Glycolipid transfer proteins (GLTPs) originally were identified as small (~24 kDa), soluble, amphitropic proteins that specifically accelerate the intermembrane transfer of glycolipids. GLTPs and related homologs now are known to adopt a unique, helically dominated, two-layer 'sandwich' architecture defined as the GLTP-fold that provides the structural underpinning for the eukaryotic GLTP superfamily. Recent advances now provide exquisite insights into structural features responsible for lipid headgroup selectivity as well as the adaptability of the hydrophobic compartment for accommodating hydrocarbon chains of differing length and unsaturation. A new understanding of the structural versatility and evolutionary premium placed on the GLTP motif has emerged. Human GLTP-motifs have evolved to function not only as glucosylceramide binding/transferring domains for phosphoinositol 4-phosphate adaptor protein-2 during glycosphingolipid biosynthesis but also as selective binding/transfer proteins for ceramide-1-phosphate. The latter, known as ceramide-1-phosphate transfer protein, recently has been shown to form GLTP-fold while critically regulating Group-IV cytoplasmic phospholipase A2 activity and pro-inflammatory eicosanoid production.
TL;DR: The unique positioning of Trp208 at the HET-C2 C terminus revealed membrane-induced conformational changes that precede glycolipid uptake, whereas key differences in residues of the sugar headgroup recognition center accounted for altered glycolIPid specificity and suggested evolutionary adaptation for the simpler glycosphingolipid compositions of filamentous fungi.
Abstract: HET-C2 is a fungal protein that transfers glycosphingolipids between membranes and has limited sequence homology with human glycolipid transfer protein (GLTP). The human GLTP fold is unique among lipid binding/transfer proteins, defining the GLTP superfamily. Herein, GLTP fold formation by HET-C2, its glycolipid transfer specificity, and the functional role(s) of its two Trp residues have been investigated. X-ray diffraction (1.9 Å) revealed a GLTP fold with all key sugar headgroup recognition residues (Asp66, Asn70, Lys73, Trp109, and His147) conserved and properly oriented for glycolipid binding. Far-UV CD showed secondary structure dominated by α-helices and a cooperative thermal unfolding transition of 49 °C, features consistent with a GLTP fold. Environmentally induced optical activity of Trp/Tyr/Phe (2:4:12) detected by near-UV CD was unaffected by membranes containing glycolipid but was slightly altered by membranes lacking glycolipid. Trp fluorescence was maximal at ∼355 nm and accessible to aqueous quenchers, indicating free exposure to the aqueous milieu and consistent with surface localization of the two Trps. Interaction with membranes lacking glycolipid triggered significant decreases in Trp emission intensity but lesser than decreases induced by membranes containing glycolipid. Binding of glycolipid (confirmed by electrospray injection mass spectrometry) resulted in a blue-shifted emission wavelength maximum (∼6 nm) permitting determination of binding affinities. The unique positioning of Trp208 at the HET-C2 C terminus revealed membrane-induced conformational changes that precede glycolipid uptake, whereas key differences in residues of the sugar headgroup recognition center accounted for altered glycolipid specificity and suggested evolutionary adaptation for the simpler glycosphingolipid compositions of filamentous fungi.
TL;DR: The self assembling properties of taurolipids were used to prepare stable copper nanoparticles (CuNPs), and the ability of CuNPs to eradicate the biofilms formed by waterborne pathogens was demonstrated.
Abstract: In this paper, the self assembling properties of taurolipids were used to prepare stable copper nanoparticles (CuNPs), and demonstrated the ability of CuNPs to eradicate the biofilms formed by waterborne pathogens. The synthesized CuNPs display wine red color and exhibited surface plasmon resonance with a maximum at 590 nm. Transmission electron microscopy showed that the CuNPs are well-dispersed with spherical morphology and the size range between 5 and 12 nm. The powder X-ray diffraction study revealed that the CuNPs was free from copper oxide impurities and crystalline with the face centered cubic structure. The CuNPs exhibited excellent anti-biofilm activity against water borne pathogens such as Escherichia coli, Pseudomonas aeruginosa, Salmonella typhi, and Shigella flexneri. Light microscopy and scanning electron microscopy (SEM) study revealed that CuNPs eliminates the mature biofilm at the minimum biofilm eradication concentration of 12.5 μM. The antimicrobial activity of the CuNPs was observed at the minimum inhibitory concentration of 25 μM, indicating the reported CuNPs exhibit true anti-biofilm effect. Fluorescence microscopy and SEM study proved that CuNPs kills the bacteria through membrane damage. The possibility to use CuNPs in cleaning biofilm formed on storage containers was demonstrated through removing the mature biofilm formed on a glass pipe.
TL;DR: The synthesis of gold nanoparticles (AuNPs) using the interdigitized vesicles formed by N-decanoyltromethamine (NDTM) was reported, and the proposed method was successfully applied for the determination of Pb2+ ions in tap water and sewage water.
Abstract: Lead is highly toxic. The detection of lead in the environmental bodies is difficult, because it is colourless and odourless. Herein, we report the synthesis of gold nanoparticles (AuNPs) using the interdigitized vesicles formed by N-decanoyltromethamine (NDTM). AuNPs stabilized by NDTM was pink in colour with spherical shape and the size is 29 ± 7 nm. The optical property of the NDTM-AuNPs was explored for the first time to detect toxic chemical, Pb2+. The addition of toxic metal ion Pb2+ to NDTM-AuNPs rapidly (
01 Jan 2016
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TL;DR: In this article, the authors present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes.
Abstract: In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
TL;DR: These results highlight critical roles for bioactive sphingolipids in most, if not all, major cell biological responses, including all major cell signalling pathways, and they link sphingoipid metabolism to key human diseases.
Abstract: Studies of bioactive lipids in general and sphingolipids in particular have intensified over the past several years, revealing an unprecedented and unanticipated complexity of the lipidome and its many functions, which rivals, if not exceeds, that of the genome or proteome. These results highlight critical roles for bioactive sphingolipids in most, if not all, major cell biological responses, including all major cell signalling pathways, and they link sphingolipid metabolism to key human diseases. Nevertheless, the fairly nascent field of bioactive sphingolipids still faces challenges in its biochemical and molecular underpinnings, including defining the molecular mechanisms of pathway and enzyme regulation, the study of lipid-protein interactions and the development of cellular probes, suitable biomarkers and therapeutic approaches.
TL;DR: The knowledge gained in this emerging field of sphingolipid metabolism will aid in the development of new therapeutic options for inflammatory disorders.
Abstract: Sphingolipids are ubiquitous building blocks of eukaryotic cell membranes. Progress in our understanding of sphingolipid metabolism, state-of-the-art sphingolipidomic approaches and animal models have generated a large body of evidence demonstrating that sphingolipid metabolites, particularly ceramide and sphingosine-1-phosphate, are signalling molecules that regulate a diverse range of cellular processes that are important in immunity, inflammation and inflammatory disorders. Recent insights into the molecular mechanisms of action of sphingolipid metabolites and new perspectives on their roles in regulating chronic inflammation have been reported. The knowledge gained in this emerging field will aid in the development of new therapeutic options for inflammatory disorders.