Peptide-oligonucleotide conjugates as nanoscale building blocks for assembly of an artificial three-helix protein mimic

Peptide-based structures can be designed to yield artificial proteins with specific folding patterns and functions. Template-based assembly of peptide units is one design option, but the use of two orthogonal self-assembly principles, oligonucleotide triple helix and a coiled coil protein domain formation have never been realized for de novo protein design. Here, we show the applicability of peptide–oligonucleotide conjugates for self-assembly of higher-ordered protein-like structures. The resulting nano-assemblies were characterized by ultraviolet-melting, gel electrophoresis, circular dichroism (CD) spectroscopy, small-angle X-ray scattering and transmission electron microscopy. These studies revealed the formation of the desired triple helix and coiled coil domains at low concentrations, while a dimer of trimers was dominating at high concentration. CD spectroscopy showed an extraordinarily high degree of α-helicity for the peptide moieties in the assemblies. The results validate the use of orthogonal self-assembly principles as a paradigm for de novo protein design

Measurement of distal intramural spread and the optimal distal resection by naked eyes after neoadjuvant radiation for rectal cancers

BACKGROUND: The safe distance between the intraoperative resection line and the visible margin of the distal rectal tumor after preoperative radiotherapy is unclear. We aimed to investigate the furthest tumor intramural spread distance in fresh tissue to determine a safe distal intraoperative resection margin length. METHODS: Twenty rectal cancer specimens were collected after preoperative radiotherapy. Tumor intramural spread distances were defined as the distance between the tumor’s visible and microscopic margins. Visible tumor margins in fresh specimens were identified during the operation and were labeled with 5 - 0 sutures under the naked eye at the distal 5, 6, and 7 o’clock directions of visible margins immediately after removal of the tumor. After fixation with formalin, the sutures were injected with nanocarbon particles. Longitudinal tissues were collected along three labels and stained with hematoxylin and eosin. The spread distance after formalin fixation was measured between the furthest intramural spread of tumor cells and the nanocarbon under a microscope. A positive intramural spread distance indicated that the furthest tumor cell was distal to the nanocarbon, and a negative value indicated that the tumor cell was proximal to the nanocarbon. The tumor intramural spread distance in fresh tissue during the operation was 1.75 times the tumor intramural spread distance after formalin fixation according to the literature. RESULTS: At the distal 5, 6, and 7 o’clock direction, seven (35%), five (25%), and six (30%) patients, respectively, had distal tumor cell intramural spread distance > 0 mm. The mean and 95% confidence interval of tumor cell intramural spread distance in fresh tissue during operation was − 0.3 (95%CI − 4.0 ~ 3.4) mm, − 0.9 (95%CI − 3.4 ~ 1.7) mm, and − 0.4 (95%CI − 3.5 ~ 2.8) mm, respectively. The maximal intraoperative intramural spread distances in fresh tissue were 8.8, 7, and 7 mm, respectively. CONCLUSIONS: The intraoperative distance between the distal resection line and the visible margin of the rectal tumor after radiotherapy should not be less than 1 cm to ensure oncological safety

Synthesis and properties of triplex-forming oligonucleotides containing 2'-modified nucleoside analogues

Triplex-forming oligonucleotides (TFOs) bind to the major groove of the DNA duplex via the Hoogsteen interactions to generate triple helices. Potential applications of triplex technology are in regulation of gene expression, site-directed gene-knockout, mutation correction and as tools in molecular biotechnology. The presence of 2’-modified nucleosides in therapeutic oligonucleotides inhibits enzymatic degradation in vivo. Therefore such sugar modifications have the potential to improve the biological activity of TFOs. We have synthesized the phosphoramidite monomers of six 2’-modified nucleosides from D-ribose via 1-O-methyl-3,5-di-O-benzyl-?-D-ribofuranoside. Three of these are N-linked nucleosides: 2’-O-methoxyethyl-5-propargylamino-uridine (MEPU), 2’-O methoxyethyl-5-methyl-cytidine (MOE-5-MeC) and 2’-O-aminoethylthymidine (AE-T); and three are C-linked nucleosides: 3-methyl-2-amino-pyrdine-2’-O-methyl-ribonucleoside (Me-MAP), 3-methyl-2- amino-pyrdine-2’-O-methoxyethyl-ribonucleoside (MOE-MAP) and 3-methyl-2- amino-pyrdine-2’-O-aminoethyl-ribonucleoside (AE-MAP). These monomers were incorporated into a number of oligonucleotides, on which the biophysical and biochemical studies have been performed. TFOs containing the N-nucleoside (MEPU) showed high duplex affinity and strong nuclease resistance. Studies on two C-nucleosides (Me-MAP and MOE-MAP) revealed that their triplex stability was determined by the sequence context. The incorporation of Me-MAP and MOE-MAP into oligonucleotides renders them much more resistant to the degradation by serum nucleobases compared to their 2’-deoxy derivative (dMAP) and 2’-deoxycytidine (dC). AE-MAP is a promising triplex stabilizer, which not only shows the highest triplex stabilization, but also displays an impressive resistance to enzymatic degradatio

Deregulation of lncRNA HIST1H2AG-6 and AIM1-3 in peripheral blood mononuclear cells is associated with newly diagnosed type 2 diabetes

Abstract Background Type 2 diabetes mellitus (T2DM) is mainly affected by genetic and environmental factors; however, the correlation of long noncoding RNAs (lncRNAs) with T2DM remains largely unknown. Methods Microarray analysis was performed to identify the differentially expressed lncRNAs and messenger RNAs (mRNAs) in patients with T2DM and healthy controls, and the expression of two candidate lncRNAs (lnc-HIST1H2AG-6 and lnc-AIM1-3) were further validated using quantitative real-time polymerase chain reaction (qRT-PCR). Spearman’s rank correlation coefficient was used to measure the degree of association between the two candidate lncRNAs and differentially expressed mRNAs. Furthermore, the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway and GO (Gene Ontology) enrichment analysis were used to reveal the biological functions of the two candidate lncRNAs. Additionally, multivariate logistic regression analysis and receiver operating characteristic (ROC) curve analysis were performed. Results The microarray analysis revealed that there were 55 lncRNAs and 36 mRNAs differentially expressed in patients with T2DM compared with healthy controls. Notably, lnc-HIST1H2AG-6 was significantly upregulated and lnc-AIM1-3 was significantly downregulated in patients with T2DM, which was validated in a large-scale qRT-PCR examination (90 controls and 100 patients with T2DM). Spearman’s rank correlation coefficient revealed that both lncRNAs were correlated with 36 differentially expressed mRNAs. Furthermore, functional enrichment (KEGG and GO) analysis demonstrated that the two lncRNA-related mRNAs might be involved in multiple biological functions, including cell programmed death, negative regulation of insulin receptor signal, and starch and sucrose metabolism. Multivariate logistic regression analysis revealed that lnc-HIST1H2AG-6 and lnc-AIM1-3 were significantly correlated with T2DM (OR = 5.791 and 0.071, respectively, both P = 0.000). Furthermore, the ROC curve showed that the expression of lnc-HIST1H2AG-6 and lnc-AIM1-3 might be used to differentiate patients with T2DM from healthy controls (area under the ROC curve = 0.664 and 0.769, respectively). Conclusion The profiles of lncRNA and mRNA were significantly changed in patients with T2DM. The expression levels of lnc-HIST1H2AG-6 and lnc-AIM1-3 genes were significantly correlated with some features of T2DM, which may be used to distinguish patients with T2DM from healthy controls and may serve as potential novel biomarkers for diagnosis in the future

Enhanced H-bonding and π-stacking in DNA: A potent duplex-stabilizing and mismatch sensing nucleobase analogue.

X-pyrene is a new nucleic acid duplex stabilizing cytosine analogue that combines enhanced π-stacking, hydrogen bonding and electrostatic interactions to greatly increase the stability of bulged DNA duplexes and DNA/RNA hybrids. X-pyrene is highly selective for guanine as a partner and duplex stability is reduced dramatically when X-pyrene or a neighboring base is mismatched. An NMR study indicates that the pyrene moiety stacks within the helix, and large changes in fluorescence emission on duplex formation are consistent with this. These favorable properties make X-pyrene a promising cytosine analogue for use in a variety of biological applications

Oligonucleotides Containing Aminated 2'-Amino-LNA Nucleotides: Synthesis and Strong Binding to Complementary DNA and RNA

Mono- and diaminated 2′-amino-LNA monomers were synthesized and introduced into oligonucleotides. Each modification imparts significant stabilization of nucleic acid duplexes and triplexes, excellent sequence selectivity, and significant nuclease resistance. Molecular modeling suggested that structural stabilization occurs via intrastrand electrostatic attraction between the protonated amino groups of the aminated 2′-amino-LNA monomers and the host oligonucleotide backbone

Causes of Recurrence of Rice Chilo suppressalis (Walker) in Longyou County and Prevention and Control measures

In recent years, the occurrence of rice Chilo suppressalis (Walker) in Longyou County of Zhejiang Province has increased year by year and has become aggravated. This poses a great threat to the safety of rice production. In order to curb the recurrence momentum of rice C. suppressalis, according to the dynamic monitoring and field system survey of rice C. suppressalis in Longyou County since 2015, the causes of the annual increase of rice C. suppressalis in recent years were analyzed, and the recurrence area of C. suppressalis was introduced, and finally some prevention and control measures were put forward

An exceptional fluorescence turn-on nucleoside: Lighting up single-stranded DNA with constant brightness regardless neighboring bases

Fluorescent nucleobase analogs (FBAs) have proven valuable for studying nucleic acid structure and dynamics. Regrettably, most FBAs exhibit reduced quantum yields when incorporated into DNA, particularly when neighboring residues are present. In this study, we introduce a turn-on nucleoside (thieno cyclopenta -dU, 3b) that increases the brightness of single-stranded oligonucleotides by approximately 10-fold compared to the free nucleoside, regardless of neighboring bases. Furthermore, an up to 50-fold increase in brightness is observed during duplex formation. To the best of our knowledge, compound 3b is the only turn-on type fluorescent nucleoside known to maintain a stable quantum yield after incorporation, and it can be well-accepted by DNA polymerases. These findings highlight the potential of turn-on FBAs for fluorescence sensing applications in enzymatic DNA synthesis and in vivo strand hybridization