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Youngbin Baek Nripen Singh Abhiram Arunkumar Ameya Borwankar Andrew L. Zydney 《Biotechnology journal》2019,14(7)
There is extensive experimental data showing that the final pH and buffer composition after protein diafiltration (DF), particularly with monoclonal antibodies, can be considerably different than that in the DF buffer due to electrostatic interactions between the charged protein and the charged ions. Previous models for this behavior have focused on the final (equilibrium) partitioning and are unable to explain the complex pH and concentration profiles during the DF process. The objective of this study is to develop a new model for antibody DF based on solution of the transient mass balance equations, with the permeate concentrations of the charged species evaluated assuming Donnan equilibrium across the semipermeable membrane in combination with electroneutrality constraints. Model predictions are in excellent agreement with experimental data obtained during DF of both acidic and basic monoclonal antibodies, with the protein charge determined from independent electrophoretic mobility measurements. The model is able to predict the entire pH/histidine concentration profiles during DF, providing a framework for the development of DF processes that yield the desired antibody formulation. 相似文献
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Ina Bock Arunkumar Dhayalan Srikanth Kudithipudi Ole Brandt Philipp Rathert Albert Jeltsch 《Epigenetics》2011,6(2):256-263
Chromatin structure is greatly influenced by histone tail post-translational modifications (PTM), which also play a central role in epigenetic processes. Antibodies against modified histone tails are central research reagents in chromatin biology and molecular epigenetics. We applied Celluspots peptide arrays for the specificity analysis of 36 commercial antibodies from different suppliers, which are directed towards modified histone tails. The arrays contained 384 peptides from eight different regions of the N-terminal tails of histones, viz. H3 1–19, 7–26, 16–35 and 26–45, H4 1–19 and 11–30, H2A 1–19 and H2B 1–19, featuring 59 post-translational modifications in many different combinations. Using various controls we document the reliability of the method. Our analysis revealed previously undocumented details in the specificity profiles of the tested antibodies. Most of the antibodies bound well to the PTM they have been raised for, but some failed. In addition, some antibodies showed high cross-reactivity and most antibodies were inhibited by specific additional PTMs close to the primary one. Furthermore, specificity profiles for antibodies directed toward the same modification sometimes were very different. The specificity of antibodies used in epigenetic research is an important issue. We provide a catalog of antibody specificity profiles for 36 widely used commercial histone tail PTM antibodies. Better knowledge about the specificity profiles of antibodies will enable researchers to implement necessary control experiments in biological studies and allow more reliable interpretation of biological experiments using these antibodies.Key words: histone modification, histone methylation, histone acetylation, histone phosphorylation, chromatin, antibody, specificity, ChIP 相似文献
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Å Västermark A Krishnan ME Houle R Fredriksson JM Cerdá-Reverter HB Schiöth 《PloS one》2012,7(7):e40982
The Agouti-like peptides including AgRP, ASIP and the teleost-specific A2 (ASIP2 and AgRP2) peptides have potent and diverse functional roles in feeding, pigmentation and background adaptation mechanisms. There are contradictory theories about the evolution of the Agouti-like peptide family as well the nomenclature. Here we performed comprehensive mining and annotation of vertebrate Agouti-like sequences. We identified A2 sequences from salmon, trout, seabass, cod, cichlid, tilapia, gilt-headed sea bream, Antarctic toothfish, rainbow smelt, common carp, channel catfish and interestingly also in lobe-finned fish. Moreover, we surprisingly found eight novel homologues from the kingdom of arthropods and three from fungi, some sharing the characteristic C-x(6)-C-C motif which are present in the Agouti-like sequences, as well as approximate sequence length (130 amino acids), positioning of the motif sequence and sharing of exon-intron structures that are similar to the other Agouti-like peptides providing further support for the common origin of these sequences. Phylogenetic analysis shows that the AgRP sequences cluster basally in the tree, suggesting that these sequences split from a cluster containing both the ASIP and the A2 sequences. We also used a novel approach to determine the statistical evidence for synteny, a sinusoidal Hough transform pattern recognition technique. Our analysis shows that the teleost AgRP2 resides in a chromosomal region that has synteny with Hsa 8, but we found no convincing synteny between the regions that A2, AgRP and ASIP reside in, which would support that the Agouti-like peptides were formed by whole genome tetraplodization events. Here we suggest that the Agouti-like peptide genes were formed through classical subsequent gene duplications where the AgRP is the most distantly related to the three other members of that group, first splitting from a common ancestor to ASIP and A2, and then later the A2 split from ASIP followed by a split resulting in ASIP2 and AgRP2. 相似文献
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Molecular phylogeny of some of the economically important silkmoths was derived using three mitochondrial genes, 12S rRNA, 16S rRNA, and COI, and the control region (CR). Maximum likelihood (ML) analyses showed two distinct clades, one consisting of moths from Bombycidae family and the other from Saturniidae family. The mitochondrial CR showed length polymorphisms with indels. The ML analyses for complete mitochondrial genome sequences of Bombyx mori (strains Aojuku, C108, Backokjam, and Xiafang), Japanese and Chinese strains of B. mandarina (Japanese mandarina and Chinese mandarina) and, Antheraea pernyi revealed two distinct clades, one comprising of B. mori strains and the other with B. mandarina, and A. pernyi forming an outgroup. Pairwise distances revealed that all of the strains of B. mori studied are closer to Chinese than to Japanese mandarina. Phylogenetic analyses based on whole mitochondrial genome sequences, the finding of a tandem triplication of a 126bp repeat element only in Japanese mandarina, and chromosome number variation in B. mandarina suggest that B. mori must have shared its recent common ancestor with Chinese mandarina. Another wild species of the Bombycidae family, Theophila religiosa, whose phylogenetic status was not clear, clustered together with the other bombycid moths in the study. Analysis of the interspecific hybrid, A. proylei gave evidence for paternal inheritance of mitochondrial DNA. 相似文献
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Thyagarajan R Arunkumar N Song W 《Journal of immunology (Baltimore, Md. : 1950)》2003,170(12):6099-6106
The B cell Ag receptor (BCR) can distinguish subtle differences in Ag structure and trigger differential responses. In this study, we analyzed the effects of Ag valency on the signaling and Ag-targeting functions of the BCR. Although both paucivalent and polyvalent Ags induced the redistribution of the surface BCR into polarized caps, polyvalent Ag-induced BCR caps persisted. Ganglioside G(M1), a lipid raft marker, and tyrosine-phosphorylated proteins, but not CD45 and transferrin receptor, were concentrated in BCR caps, suggesting BCR caps as surface-signaling microdomains. Prolonged BCR caps were concomitant with an increase in the level and duration of protein tyrosine phosphorylation and a reduction in BCR internalization and movement to late endosomes/lysosomes. Thus, Ag valency influences B cell responses by modulating the stability of BCR-signaling microdomains and BCR trafficking. 相似文献
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Saikia Banashree Singh Sanjay Debbarma Johni Velmurugan Natarajan Dekaboruah Hariprasanna Arunkumar Kallare P. Chikkaputtaiah Channakeshavaiah 《Physiology and Molecular Biology of Plants》2020,26(5):857-869
Physiology and Molecular Biology of Plants - The recent global climate change has directly impacted major biotic and abiotic stress factors affecting crop productivity worldwide. Therefore, the... 相似文献
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Arunkumar Venkatesan Jie Geng Malathi Kandarpa Sanjeeva Joseph Wijeyesakere Ashwini Bhide Moshe Talpaz Irina D. Pogozheva Malini Raghavan 《The Journal of cell biology》2021,220(7)
Myeloproliferative neoplasms (MPNs) are frequently driven by mutations within the C-terminal domain (C-domain) of calreticulin (CRT). CRTDel52 and CRTIns5 are recurrent mutations. Oncogenic transformation requires both mutated CRT and the thrombopoietin receptor (Mpl), but the molecular mechanism of CRT-mediated constitutive activation of Mpl is unknown. We show that the acquired C-domain of CRTDel52 mediates both Mpl binding and disulfide-linked CRTDel52 dimerization. Cysteine mutations within the novel C-domain (C400A and C404A) and the conserved N-terminal domain (N-domain; C163A) of CRTDel52 are required to reduce disulfide-mediated dimers and multimers of CRTDel52. Based on these data and published structures of CRT oligomers, we identify an N-domain dimerization interface relevant to both WT CRT and CRTDel52. Elimination of disulfide bonds and ionic interactions at both N-domain and C-domain dimerization interfaces is required to abrogate the ability of CRTDel52 to mediate cell proliferation via Mpl. Thus, MPNs exploit a natural dimerization interface of CRT combined with C-domain gain of function to achieve cell transformation. 相似文献
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Johni Debbarma Banashree Saikia Dhanawantari L. Singha Jitendra Maharana Natarajan Velmuruagan Hariprasanna Dekaboruah Kallare P. Arunkumar Channakeshavaiah Chikkaputtaiah 《Physiology and Molecular Biology of Plants》2021,27(7):1559
Fusarium wilt caused by Fusarium oxysporum f. sp. lycopersici (Fol) is a major fungal disease of tomato (Solanum lycopersicum L.). Xylem sap protein 10 (XSP10) and Salicylic acid methyl transferase (SlSAMT) have been identified as putative negative regulatory genes associated with Fusarium wilt of tomato. Despite their importance as potential genes for developing Fusarium wilt disease tolerance, very little knowledge is available about their expression, cell biology, and functional genomics. Semi-quantitative and quantitative real-time PCR expression analysis of XSP10 and SlSAMT, in this study, revealed higher expression in root and flower tissue respectively in different tomato cultivars viz. Micro-Tom (MT), Arka Vikas (AV), and Arka Abhed (AA). Therefore, the highly up-regulated expression of XSP10 and SlSAMT in biotic stress susceptible tomato cultivar (AV) than a multiple disease resistant cultivar (AA) suggested the disease susceptibility nature of these genes for Fusarium wilt. Sub-cellular localization analysis through the expression of gateway cloning constructs in tomato protoplasts and seedlings showed the predominant localization of XSP10 in the nucleus and SlSAMT at the cytoplasm. A strong in vivo protein–protein interaction of XSP10 with SlSAMT at cytoplasm from bi-molecular fluorescent complementation study suggested that these two proteins function together in regulating responses to Fusarium wilt tolerance in tomato.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12298-021-01025-y. 相似文献