63 research outputs found
Dissimilarity in the Folding of Human Cytosolic Creatine Kinase Isoenzymes
Creatine kinase (CK, EC 2.7.3.2) plays a key role in the energy homeostasis of excitable cells. The cytosolic human CK isoenzymes exist as homodimers (HMCK and HBCK) or a heterodimer (MBCK) formed by the muscle CK subunit (M) and/or brain CK subunit (B) with highly conserved three-dimensional structures composed of a small N-terminal domain (NTD) and a large C-terminal domain (CTD). The isoforms of CK provide a novel system to investigate the sequence/structural determinants of multimeric/multidomain protein folding. In this research, the role of NTD and CTD as well as the domain interactions in CK folding was investigated by comparing the equilibrium and kinetic folding parameters of HMCK, HBCK, MBCK and two domain-swapped chimeric forms (BnMc and MnBc). Spectroscopic results indicated that the five proteins had distinct structural features depending on the domain organizations. MBCK BnMc had the smallest CD signals and the lowest stability against guanidine chloride-induced denaturation. During the biphasic kinetic refolding, three proteins (HMCK, BnMc and MnBc), which contained either the NTD or CTD of the M subunit and similar microenvironments of the Trp fluorophores, refolded about 10-fold faster than HBCK for both the fast and slow phase. The fast folding of these three proteins led to an accumulation of the aggregation-prone intermediate and slowed down the reactivation rate thereby during the kinetic refolding. Our results suggested that the intra- and inter-subunit domain interactions modified the behavior of kinetic refolding. The alternation of domain interactions based on isoenzymes also provides a valuable strategy to improve the properties of multidomain enzymes in biotechnology
Cross-Regulation between Oncogenic BRAFV600E Kinase and the MST1 Pathway in Papillary Thyroid Carcinoma
BACKGROUND:The BRAF(V600E) mutation leading to constitutive signaling of MEK-ERK pathways causes papillary thyroid cancer (PTC). Ras association domain family 1A (RASSF1A), which is an important regulator of MST1 tumor suppressor pathways, is inactivated by hypermethylation of its promoter region in 20 to 32% of PTC. However, in PTC without RASSF1A methylation, the regulatory mechanisms of RASSF1A-MST1 pathways remain to be elucidated, and the functional cooperation or cross regulation between BRAF(V600E) and MST1,which activates Foxo3,has not been investigated. METHODOLOGY/PRINCIPAL FINDINGS:The negative regulators of the cell cycle, p21 and p27, are strongly induced by transcriptional activation of FoxO3 in BRAF(V600E) positive thyroid cancer cells. The FoxO3 transactivation is augmented by RASSF1A and the MST1 signaling pathway. Interestingly, introduction of BRAF(V600E)markedly abolished FoxO3 transactivation and resulted in the suppression of p21 and p27 expression. The suppression of FoxO3 transactivation by BRAF(V600E)is strongly increased by coexpression of MST1 but it is not observed in the cells in which MST1, but not MST2,is silenced. Mechanistically, BRAF(V600E)was able to bind to the C-terminal region of MST1 and resulted in the suppression of MST1 kinase activities. The induction of the G1-checkpoint CDK inhibitors, p21 and p27,by the RASSF1A-MST1-FoxO3 pathway facilitates cellular apoptosis, whereas addition of BRAF(V600E) inhibits the apoptotic processes through the inactivation of MST1. Transgenic induction of BRAF(V600E)in the thyroid gland results in cancers resembling human papillary thyroid cancers. The development of BRAF(V600E)transgenic mice with the MST1 knockout background showed that these mice had abundant foci of poorly differentiated carcinomas and large areas without follicular architecture or colloid formation. CONCLUSIONS/SIGNIFICANCE:The results of this study revealed that the oncogenic effect of BRAF(V600E) is associated with the inhibition of MST1 tumor suppressor pathways, and that the activity of RASSF1A-MST1-FoxO3 pathways determines the phenotypes of BRAF(V600E) tumors
A new strategy for enhancing imputation quality of rare variants from next-generation sequencing data via combining SNP and exome chip data
Identification and quantification of glucosinolates in Korean leaf mustard germplasm (Brassica juncea var. integrifolia) by liquid chromatography–electrospray ionization/tandem mass spectrometry
Neuronal mechanisms and circuits underlying repetitive behaviors in mouse models of autism spectrum disorder
Stress update algorithm for enhanced homogeneous anisotropic hardening model
A stress integration algorithm is provided for a novel homogeneous-yield-function-based anisotropic hardening (HAH) model. The new model is an extension of the original HAH model that describes cross-hardening or softening of a sheet metal under an orthogonal strain path change. A semi-explicit integration scheme for the stress update is utilized to efficiently handle the gradient of the distorted yield surface during complex strain path changes, as originally proposed by Lee et al. (2012). Validations of the algorithm developed were conducted by comparing the predicted stress-strain curves of dual-phase (DP) 780 and extra-deep-drawing-quality (EDDQ) steels with experimental stress-strain responses observed under cross-loading conditions. Finally, the accuracy of the proposed finite element (FE) formulations was assessed by r-value prediction and preparation of iso-error maps. (C) 2014 Elsevier B.V. All rights reserved.X111010sciescopu
Spatial variability in hydrogen and oxygen isotopic composition of Korean Red Pine and its implication for tracing wood origin
Rational modification of substrate binding site by structure-based engineering of a cellobiose 2-epimerase in Caldicellulosiruptor saccharolyticus
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