Self-recognition of high-mannose type glycans mediating adhesion of embryonal fibroblasts

High-mannose type N-linked glycan with 6 mannosyl residues, termed "M6Gn2", displayed clear binding to the same M6Gn2, conjugated with ceramide mimetic (cer-m) and incorporated in liposome, or coated on polystyrene plates. However, the conjugate of M6Gn2-cer-m did not interact with complex-type N-linked glycan with various structures having multiple GlcNAc termini, conjugated with cer-m. The following observations indicate that hamster embryonic fibroblast NIL-2 K cells display homotypic autoadhesion, mediated through the self-recognition capability of high-mannose type glycans expressed on these cells: (i) NIL-2 K cells display clear binding to lectins capable of binding to high-mannose type glycans (e.g., ConA), but not to other lectins capable of binding to other carbohydrates (e.g. GS-II). (ii) NIL-2 K cells adhere strongly to plates coated with M6Gn2-cer-m, but not to plates coated with complex-type N-linked glycans having multiple GlcNAc termini, conjugated with cer-m; (iii) degree of NIL-2 K cell adhesion to plates coated with M6Gn2-cer-m showed a clear dose-dependence on the amount of M6Gn2-cer-m; and (iv) the degree of NIL-2 K adhesion to plates coated with M6Gn2-cer-m was inhibited in a dose-dependent manner by α1,4-L-mannonolactone, the specific inhibitor in high-mannose type glycans addition. These data indicate that adhesion of NIL-2 K is mediated by self-aggregation of high mannose type glycan. Further studies are to be addressed on auto-adhesion of other types of cells based on self interaction of high mannose type glycans.     

Hypoglycemic dipeptide cyclo (His-Pro) significantly altered plasma proteome in streptozocin-induced diabetic rats and genetically-diabetic (ob/ob) mice

The proteins in plasma perform many important functions in the body, and the protein profiles of the plasma vary under different physiological and pathological conditions. In an attempt to identify novel marker proteins for diabetes prognosis, we examined the effect of hypoglycemic dipeptide cyclo (His-Pro) (CHP) on the differential regulation of plasma proteins in streptozocin-induced diabetic rats and genetically-diabetic (ob/ob) mice. The orally-administrated CHP produced an excellent hypoglycemic effect in both animal models, lowering the average plasma glucose level by over 50 %. In the 2-DE analysis of the plasma, a total of 23 spots among 500 visualized spots were found to be differentially regulated, and they were identified by MALDI/TOF mass spectrometry. These proteins include the down-regulation of Apo E and the up-regulation of FGA, Apo A-I, Apo A-IV, and A1M in STZ-induced diabetic rats. Moreover, CHP significantly reduced the plasma protein levels of FGB, FGC, F12, C1QTNF5, and SPA3K, as well as increased the abundance of A1M, A2M, Apo E, and TTR in genetically-diabetic mice. In conclusion, alteration in the regulation of these proteins indicates that this treatment may be successful in overcoming the diabetic state. The present proteomic data can serve as the basis for the development of specific evidence-based interventions allowing for the prevention and treatment of diabetes.     

Functional analysis and tissue-differential expression of four FAD2 genes in amphidiploid Brassica napus derived from Brassica rapa and Brassica oleracea

Fatty acid desaturase 2 (FAD2), which resides in the endoplasmic reticulum (ER), plays a crucial role in producing linoleic acid (18:2) through catalyzing the desaturation of oleic acid (18:1) by double bond formation at the delta 12 position. FAD2 catalyzes the first step needed for the production of polyunsaturated fatty acids found in the glycerolipids of cell membranes and the triacylglycerols in seeds. In this study, four FAD2 genes from amphidiploid Brassica napus genome were isolated by PCR amplification, with their enzymatic functions predicted by sequence analysis of the cDNAs. Fatty acid analysis of budding yeast transformed with each of the FAD2 genes showed that whereas BnFAD2-1, BnFAD2-2, and BnFAD2-4 are functional enzymes, and BnFAD2-3 is nonfunctional. The four FAD2 genes of B. napus originated from synthetic hybridization of its diploid progenitors Brassica rapa and Brassica oleracea, each of which has two FAD2 genes identical to those of B. napus. The BnFAD2-3 gene of B. napus, a nonfunctional pseudogene mutated by multiple nucleotide deletions and insertions, was inherited from B. rapa. All BnFAD2 isozymes except BnFAD2-3 localized to the ER. Nonfunctional BnFAD2-3 localized to the nucleus and chloroplasts. Four BnFAD2 genes can be classified on the basis of their expression patterns.     

Spotted fever group rickettsia closely related to Rickettsia monacensis isolated from ticks in South Jeolla province,Korea

Rickettsia monacensis, a spotted fever group rickettsia, was isolated from Ixodes nipponensis ticks collected from live‐captured small mammals in South Jeolla province, Korea in 2006. Homogenates of tick tissues were inoculated into L929 and Vero cell monolayers using shell vial assays. After several passages, Giemsa staining revealed rickettsia‐like organisms in the inoculated Vero cells, but not the L929 cells. Sequencing analysis revealed that the ompA‐small part (25–614 bp region), ompA‐large part (2849–4455 bp region), nearly full‐length ompB (58–4889 bp region) and gltA (196–1236 bp region) of the isolates had similarities of 100%, 99.8%, 99.3% and 99.5%, respectively, to those of R. monacensis. Furthermore, phylogenetic analysis showed that the isolate was grouped into the cluster in the same way as R. monacensis in the trees of all genes examined. These results strongly suggest that the isolate is closely related to R. monacensis. As far as is known, this is the first report of isolation of R. monacensis from ticks in Korea.     

Crystal structure of human cytosolic aspartyl‐tRNA synthetase,a component of multi‐tRNA synthetase complex

Human cytosolic aspartyl‐tRNA synthetase (DRS) catalyzes the attachment of the amino acid aspartic acid to its cognate tRNA and it is a component of the multi‐tRNA synthetase complex (MSC) which has been known to be involved in unexpected signaling pathways. Here, we report the crystal structure of DRS at a resolution of 2.25 Å. DRS is a homodimer with a dimer interface of 3750.5 Ų which comprises 16.6% of the monomeric surface area. Our structure reveals the C‐terminal end of the N‐helix which is considered as a unique addition in DRS, and its conformation further supports the switching model of the N‐helix for the transfer of tRNA^Asp to elongation factor 1α. From our analyses of the crystal structure and post‐translational modification of DRS, we suggest that the phosphorylation of Ser146 provokes the separation of DRS from the MSC and provides the binding site for an interaction partner with unforeseen functions.Proteins 2013; 81:1840–1846. © 2013 Wiley Periodicals, Inc.     

The structure of a shellfish specific GST class glutathione S‐transferase from antarctic bivalve Laternula elliptica reveals novel active site architecture

Glutathione‐S‐transferases have been identified in all the living species examined so far, yet little is known about their function in marine organisms. In a previous report, the recently identified GST from Antarctic bivalve Laternula elliptica (LeGST) was classified into the rho class GST, but there are several unique features of LeGST that may justify reclassification, which could represent specific shellfish GSTs. Here, we determined the crystal structure of LeGST, which is a shellfish specific class of GST. The structural analysis showed that the relatively open and wide hydrophobic H‐site of the LeGST allows this GST to accommodate various substrates. These results suggest that the H‐site of LeGST may be the result of adaptation to their environments as sedentary organisms. Proteins 2013. © 2012 Wiley Periodicals, Inc.