An efficient protocol for calculating 13C NMR chemical
shifts for natural products with multiple degrees of conformational
freedom is described. This involves a multistep procedure starting
from molecular mechanics and ending with a large basis set density
functional model to obtain accurate Boltzmann conformer weights, followed
by empirically corrected density functional NMR calculations for the
individual conformers. The accuracy of the protocol (average rms <4
ppm) was determined by application to ∼925 diverse natural
products, the structures of which have been confirmed either by X-ray
crystallography or independent synthesis. The protocol was then applied
to ∼ 2275 natural products, the structures of which were elucidated
mainly by NMR and MS data. Five to ten percent of the latter compounds
exhibited rms errors significantly in excess of 4 ppm, suggesting
possible structural or signal assignment errors. Both data sets are
available from an online browser (). The procedure can be and has been fully automated
and is practical using present-generation personal computers, requiring
a few hours or days depending on the size of the molecule and number
of accessible conformers.
Tumor necrosis factor (TNF)-alpha is one of the candidate mediators of insulin resistance associated with obesity, a major risk factor for the development of type 2 diabetes. The insulin resistance induced by TNF-alpha is antagonized by thiazolidinediones (TZDs), a new class of insulin-sensitizing drugs. The aim of the current study was to dissect the mechanism whereby pioglitazone, one of the TZDs, ameliorates TNF-alpha-induced insulin resistance in 3T3-L1 adipocytes. Pioglitazone restored insulin-stimulated 2-deoxyglucose (DOG) uptake, which was reduced by TNF-alpha, with concomitant restorations in tyrosine phosphorylation and protein levels of insulin receptor (IR) and insulin receptor substrate (IRS)-1, as well as association of the p85 regulatory subunit of phosphatidylinositol (PI) 3-kinase with IRS-1 and PI 3-kinase activity. Adenovirus-mediated gene transfer of either wild-type human peroxisome proliferator-activated receptor (PPAR)-gamma2 or a mutant carrying a replacement at the consensus mitogen-activated protein kinase phosphorylation site (hPPAR-gamma2-S112A) promoted adipogenesis of 3T3-L1 fibroblasts and restored TNF-alpha-induced decrease of triglyceride in adipocytes as effectively as pioglitazone. Overexpression of the PPAR-gamma proteins in TNF-alpha-treated adipocytes restored protein levels of IR/IRS-1, but did not improve insulin-stimulated tyrosine phosphorylation of IR/IRS-1 or insulin-stimulated 2-DOG uptake. These results indicate that the ability of pioglitazone to restore insulin-stimulated tyrosine phosphorylation of IR/IRS-1, which is necessary for amelioration of TNF-alpha-induced insulin resistance, may be independent of the adipogenic activity of PPAR-gamma that regulates protein levels of IR/IRS-1.
We previously reported three families with type A insulin-resistant syndrome who had mutations, either Asp 1179 or Leu 1193 , in the kinase domain of the insulin receptor. The extreme insulin resistance of these patients was found to be caused by the decreased number of insulin receptors on the cell surface, due to the intracellular rapid degradation (Imamura, T.,
The Raney nickel catalyst was employed as a catalytic electrode in the electrohydrogenation of various unsaturated compounds, including ketones, aromatic aldehydes, Schiff’s bases, oximes, nitriles, aromatic nitro compounds, olefins, and acetylenes. The hydrogen generated electrochemically was adsorbed and activated at the surface of the catalyst; it effectively hydrogenated these compounds, affording products which are very similar to those obtained in a normal catalytic hydrogenation with Raney nickel.
In some microorganisms, polyunsaturated fatty acids (PUFAs) are biosynthesized by PUFA synthases characterized by tandem acyl carrier proteins (ACPs) in subunit A. These ACPs were previously shown to be important for PUFA productivity. In this study, we examined their function in more detail. PUFA productivities increased depending on the number of ACPs without profile changes in each subunit A of eukaryotic and prokaryotic PUFA synthases. We also constructed derivative enzymes from subunit A with 5 × ACPs. Enzymes possessing one inactive ACP at any position produced ~30% PUFAs compared with the parental enzyme but unexpectedly had ~250% productivity compared with subunit A with 4 × ACPs. Enzymes constructed by replacing the 3rd ACP with an inactive ACP from another subunit A or ACP-unrelated sequences produced ~100% and ~3% PUFAs compared with the parental 3rd ACP-inactive enzyme, respectively. These results suggest that both the structure and number of ACP domains are important for PUFA productivity.
Shc has two distinct domains, amino-terminal and SH2 domain, which can interact with activated growth factor receptors. Shc interacts with insulin receptor via Shc-amino-terminal (N) domain, whereas Shc associates with epidermal growth factor (EGF) receptor through both Shc-N and -SH2 domains. In accordance with the different functional roles between insulin and EGF receptors, EGF stimulated tyrosine phosphorylation of Shc faster than insulin. To clarify the functional importance of three distinct Shc domains on insulin and EGF signaling, we microinjected glutathione S-transferase (GST) fusion proteins containing the amino terminus plus collagen homology domain (NCH), collagen homology domain (CH), and Src homology 2 domain (SH2) into Rat1 fibroblasts expressing insulin receptors (HIRc). Bromodeoxyuridine (BrdUrd) incorporation into newly synthesized DNA was subsequently studied to assess the importance of the three distinct domains of Shc. Microinjection of the NCH-GST fusion protein inhibited BrdUrd incorporation induced by both EGF and insulin, whereas microinjection of the SH2-GST fusion protein inhibited EGF, but not insulin stimulation of DNA synthesis. Neither EGF-nor insulin-induced BrdUrd incorporation was inhibited by the CH-GST fusion protein.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.