Solvent effects on the stability of A7U7p

The thermodynamics of double-helix formation were measured spectrophotometrically for A7U7 in water at 1 M NaCl and for A7U7p in a variety of solvent mixtures and salt. Comparison of the A7U7 results with calorimetric measurements indicates duplex formation involves intermediate states. For A7U7p between 0.06 and 0.55 M Na+, dTm/d(log [Na+]) = 17.4 degrees C, similar to the value of 19.6 degrees C for poly-(A).poly(U) [Krakauer, H., & Sturtevant, J. M. (1968) Biopolymers 6, 491-512]. At 1 M NaCl, the A7U7p duplex is most stable in 100% water. For 10 mol % solutions, the order for A7U7p duplex stability is ethylene glycol greater than glycerol greater than ethanol greater than 2-propanol greater than dimethyl sulfoxide greater than 1-propanol greater than formamide greater than N,N-dimethylformamide greater than urea greater than dioxane. Comparison of changes in stability and thermodynamic parameters with literature results for proteins suggests proteins and A7U7p interact differently with solvent. The results suggest hydrophobic bonding is not a major contributor to the stability of the A7U7p duplex. Comparisons with bulk solvent surface tension suggest the energy of cavity formation is also not a major contributor to duplex stability.     

Evolution of the B7 family: co-evolution of B7H6 and NKp30, identification of a new B7 family member, B7H7, and of B7's historical relationship with the MHC

The B7 family of genes is essential in the regulation of the adaptive immune system. Most B7 family members contain both variable (V)- and constant (C)-type domains of the immunoglobulin superfamily (IgSF). Through in silico screening of the Xenopus genome and subsequent phylogenetic analysis, we found novel genes belonging to the B7 family, one of which is the recently discovered B7H6. Humans and rats have a single B7H6 gene; however, many B7H6 genes were detected in a single large cluster in the Xenopus genome. The B7H6 expression patterns also varied in a species-specific manner. Human B7H6 binds to the activating natural killer receptor, NKp30. While the NKp30 gene is single-copy and maps to the MHC in most vertebrates, many Xenopus NKp30 genes were found in a cluster on a separate chromosome that does not harbor the MHC. Indeed, in all species so far analyzed from sharks to mammals, the number of NKp30 and B7H6 genes correlates well, suggestive of receptor-ligand co-evolution. Furthermore, we identified a Xenopus-specific B7 homolog (B7HXen) and revealed its close linkage to B2M, which we have demonstrated previously to have been originally encoded in the MHC. Thus, our study provides further proof that the B7 precursor was included in the proto MHC. Additionally, the comparative analysis revealed a new B7 family member, B7H7, which was previously designated in the literature as an unknown gene, HHLA2.     

Immune Checkpoints of the B7 Family. Part 2. Representatives of the B7 Family B7-H3, B7-H4, B7-H5, B7-H6, B7-H7, and ILDR2 and Their Receptors

Russian Journal of Bioorganic Chemistry - Immune checkpoints regulate polarity, strength, and termination of the immune response. The leading roles in these processes are played by molecules of the...     

Solid-phase synthesis of the self-complementary hexamer d(c7GpCpc7GpCpc7GpC) via the O-3''-phosphoramidite of 7-deaza-2''-deoxyguanosine.

The synthesis of the O-3'-phosphoramidite of a suitably protected 7-deaza-2'-deoxyguanosine (c7G) which is an isostere of 2'-deoxyguanosine is described. The phosphoramidite of the modified nucleoside was used in the synthesis of the self-complementary hexamer d(c7GpCpc7GpCpc7GpC) on functionalized silica gel in a mini-reactor. As expected from the parent hexamer d(GpCpGpCpGpC) the isosteric d(c7GpCpc7GpCpc7GpC) exhibits a rigid secondary structure (22% hypochromicity at 280 nm) and forms a duplex in 1 M aqueous sodium chloride solution. Due to the altered pi-electron system of the pyrrolo[2,3-d]pyrimidine nucleobase, which affects base stacking and hydrogen bonding, the Tm of the modified duplex is decreased by 10 degrees C compared to that of the parent purine hexamer. Moreover, it is expected that the incorporation of c7G influences the pitch of the helix.     

The prodomain of BMP-7 targets the BMP-7 complex to the extracellular matrix

Biochemical and biophysical methods are used to show that BMP-7 is secreted as a stable complex consisting of the processed growth factor dimer noncovalently associated with its two prodomain propeptide chains and that the BMP-7 complex is structurally similar to the small transforming growth factor beta (TGFbeta) complex. Because the prodomain of TGFbeta interacts with latent TGFbeta-binding proteins, a family of molecules homologous to the fibrillins, the prodomain of BMP-7 was tested for binding to fibrillin-1 or to LTBP-1. The BMP-7 prodomain and BMP-7 complex, but not the separated growth factor dimer, interact with N-terminal regions of fibrillin-1. This interaction may target the BMP-7 complex to fibrillin microfibrils in the extracellular matrix. Immunolocalization of BMP-7 in tissues like the kidney capsule and skin reveals co-localization with fibrillin. However, BMP-7 immunolocalization in other tissues known to be active sites for BMP-7 signaling is not apparent, suggesting that immunolocalization of BMP-7 in certain tissues represents specific extracellular storage sites. These studies suggest that the prodomains of TGFbeta-like growth factors are important for positioning and concentrating growth factors in the extracellular matrix. In addition, they raise the possibility that prodomains of other TGFbeta-like growth factors interact with fibrillins and/or LTBPs and are also targeted to the extracellular matrix.     

Oligonucleotide duplex stability controlled by the 7-substituents of 7-deazaguanine bases

Oligonucleotides containing 7-substituted 7-deazaguanine residues (7-methyl, 7-iodo) have been synthesized. The self-complemetary octamer d(I⁷c⁷G-C)₄ containing 7-iodo-7-deaza-2′-deoxyguanosine forms a stabilized duplex compared to the parent oligomer d(G-C)₄ (ΔT_m = +10° C). Also the complex between the oligodeoxynucleotide d(I⁷c⁷G₅-G) and poly(C) is stabilized (ΔT_m = +10°) over that of d(c⁷G₅-G) with poly(C).     

Linkage relationships of peptidase-7, Pep-7, in the mouse

An electrophoretically detectable variant of peptidase-7 in Mus musculus has been found and used to locate the structural gene, Pep-7, on chromosome 5. Gene order and recombination frequencies are estimated as Pep-7 3.5±2.0 Rw 8.8±2.2 go 20.0±4.6 bf.     

Fibroblast cell shape and adhesion in vitro is altered by overexpression of the 7a and 7b isoforms of protocadherin 7, but not the 7c isoform

Protocadherins (Pcdhs) are a family of cadherins considered to play an important role in the cell-cell adhesion of specific neurons in the central nervous system. Of the reported Pcdhs, relatively little is known about the functional role of protocadherin 7 (Pcdh7), and there is no evidence of Pcdh7 mediated cell-cell adhesion. To date, three splicing variants are known; they may have different effects on cell phenotype. We report here that mouse fibroblast L cells stably overexpressing the Pcdh7 isoforms 7a and 7b, but not 7c, showed a morphological change and Ca(2+)dependent cell adhesion.     

Amino Acid Substitutions Improve the Immunogenicity of H7N7HA Protein and Protect Mice against Lethal H7N7 Viral Challenge

Avian influenza A H7N7/NL/219/03 virus creates a serious pandemic threat to human health because it can transmit directly from domestic poultry to humans and from human to human. Our previous vaccine study reported that mice when immunized intranasally (i.n) with live Bac-HA were protected from lethal H7N7/NL/219/03 challenge, whereas incomplete protection was obtained when administered subcutaneously (s.c) due to the fact that H7N7 is a poor inducer of neutralizing antibodies. Interestingly, our recent vaccine studies reported that mice when vaccinated subcutaneously with Bac-HA (H7N9) was protected against both H7N9 (A/Sh2/2013) and H7N7 virus challenge. HA1 region of both H7N7 and H7N9 viruses are differ at 15 amino acid positions. Among those, we selected three amino acid positions (T143, T198 and I211) in HA1 region of H7N7. These amino acids are located within or near the receptor binding site. Following the selection, we substituted the amino acid at these three positions with amino acids found on H7N9HA wild-type. In this study, we evaluate the impact of amino acid substitutions in the H7N7 HA-protein on the immunogenicity. We generated six mutant constructs from wild-type influenza H7N7HA cDNA by site directed mutagenesis, and individually expressed mutant HA protein on the surface of baculovirus (Bac-HA_m) and compared their protective efficacy of the vaccines with Bac-H7N7HA wild-type (Bac-HA) by lethal H7N7 viral challenge in a mouse model. We found that mice immunized subcutaneously with Bac-HA_m constructs T143A or T198A-I211V or I211V-T143A serum showed significantly higher hemagglutination inhibition and neutralization titer against H7N7 and H7N9 viruses when compared to Bac-HA vaccinated mice groups. We also observed low level of lung viral titer, negligible weight loss and complete protection against lethal H7N7 viral challenge. Our results indicated that amino acid substitution at position 143 or 211 improve immunogenicity of H7N7HA vaccine against H7N7/NL/219/03 virus.     

Microsequence analysis of DNA-binding proteins 7a, 7b, and 7e from the archaebacterium Sulfolobus acidocaldarius

DNA-binding proteins in eubacteria, such as Escherichia coli NS1 and NS2, are generally small basic molecules. In contrast, the archaebacterium Sulfolobus acidocaldarius contains three groups of DNA-binding proteins which have molecular masses of 7, 8, and 10 kDa. In the first group, five proteins (7a-7e) have been identified, while in the second and third group only two proteins each are present, denoted 8a and 8b and 10a and 10b, respectively. In this paper, we present the primary structures of proteins 7a, 7b, and 7e from the first group. All three proteins contain lysyl residues which are monomethylated to different extents. The modified lysines are found in the NH2-terminal regions of all 7-kDa proteins and in the COOH-terminal part of protein 7e. The sequences of the 7-kDa group are highly similar to each other. All of these macromolecules have been shown to interact specifically with DNA. Protein 7e of the 7-kDa group shows the tightest binding to DNA.     

Synthesis of 7alpha-hydroxy-dehydroepiandrosterone and 7beta-hydroxy-dehydroepiandrosterone

The fermentation of dehydroepiandrosterone synthesized from the starting material diosgenin using Mucor racemosus produced 7alpha-hydroxy-dehydroepiandrosterone and 7beta-hydroxy-dehydroepiandrosterone. The bioactivity of the microbial metabolites is also discussed. The species M. racemosus was isolated by screening among stains from soil samples collected from various parts of China.