Specific Molecular Size Research Articles

Exploring the role of tRNA-derived small RNAs (tsRNAs) in disease: implications for HIF-1 pathway modulation.

The tRNA-derived small RNAs (tsRNAs) can be categorized into two main groups: tRNA-derived fragments (tRFs) and tRNA-derived stress-induced RNAs (tiRNAs). Each group possesses specific molecular sizes, nucleotide compositions, and distinct physiological functions. Notably, hypoxia-inducible factor-1 (HIF-1), a transcriptional activator dependent on oxygen, comprises one HIF-1β subunit and one HIF-α subunit (HIF-1α/HIF-2α/HIF-3α). The activation of HIF-1 plays a crucial role in gene transcription, influencing key aspects of cancer biology such as angiogenesis, cell survival, glucose metabolism, and invasion. The involvement of HIF-1α activation has been demonstrated in numerous human diseases, particularly cancer, making HIF-1 an attractive target for potential disease treatments. Through a series of experiments, researchers have identified two tiRNAs that interact with the HIF-1 pathway, impacting disease development: 5'tiRNA-His-GTG in colorectal cancer (CRC) and tiRNA-Val in diabetic retinopathy (DR). Specifically, 5'tiRNA-His-GTG promotes CRC development by targeting LATS2, while tiRNA-Val inhibits Sirt1, leading to HIF-1α accumulation and promoting DR development. Clinical data have further indicated that certain tsRNAs' expression levels are associated with the prognosis and pathological features of CRC patients. In CRC tumor tissues, the expression level of 5'tiRNA-His-GTG is significantly higher compared to normal tissues, and it shows a positive correlation with tumor size. Additionally, KEGG analysis has revealed multiple tRFs involved in regulating the HIF-1 pathway, including tRF-Val-AAC-016 in diabetic foot ulcers (DFU) and tRF-1001 in pathological ocular angiogenesis. This comprehensive article reviews the biological functions and mechanisms of tsRNAs related to the HIF-1 pathway in diseases, providing a promising direction for subsequent translational medicine research.

Molecular size distribution in pentavalent (A, C, Y, W, X) meningococcal polysaccharide conjugate vaccine by HPSEC-UV-MALS-RI method- a conceivable stability indicating parameter

Molecular size distribution (MSD) of polysaccharides serves as a key parameter that directly correlates to the immunogenicity of vaccine. MSD at meningococcal polysaccharide (A, C, Y and W) or conjugate bulk level is well established under detailed pharmacopeial and WHO guidelines. We report here, a newly developed method for determination of molecular size distribution of pentavalent Meningococcal conjugate vaccine comprising of A, C, Y, W and X (MenFive). Although serogroup specific molecular size could not be estimated here; lot to lot consistency monitoring, molecular aggregates distribution in final lot, are key takeaways of this method. Determination of MSD in pentavalent fill finished product was quite challenging. Various columns/detectors combination, buffers, physico-chemical conditions (temperature, 2–8 °C, 25 °C, 40 °C and 60 °C; flow rate, 0.3 mL to 0.8 mL), liquid/lyophilized formulations, were explored. Polymer-based packed columns were explored for estimation for MSD by aqueous size exclusion chromatography, using combinations of- Shodex OHPAK SB 807 HQ, Shodex OHPAK SB 806 HQ, G6000 PWXL, coupled with guard Shodex OHPAK SB-G-6B. MenFive showed heterogenous distribution of molecules ranging from 200 to 19000 kDa, indicating its complex nature. However, 1000–8000 kDa was dominant range, comprising of ≥ 50 % distribution of molecules, in both liquid as well as lyophilized formulations, with average molecular weight around 6000–6500 kDa. The molar mass distribution after slicing would provide an insight to the conformation of molecules through its presentation as HMW, LMW, aggregates and subsequently, the presence of dominant population of molecules of a particular molecular weight and its total contribution in the sample.

"Preparation, Fractionation and Determination of Molecular Weight of COS using MALDI-TOF-mass Spectrometry Analysis"

Chitooligosaccharides (COS) derived from the hydrolysis of high molecular weight chitosan (HMWC) was prepared and fractionated in salting-out method with a wide range of NaCl, and characterized by Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF) mass spectrometry analysis. It was confirmed that the COS before salting-out had a range from (GlcN)₄ to (GlcN)₁₅. A large decrease in COS molecules was confirmed according to the sequential salting-out, and finally, the COS according to the quaternary salting-out was mainly confirmed in (GlcN)_5~7. Unfortunately, it was not possible to efficiently separate or purify molecules larger than (GlcN)₇ in a single molecule according to salting-out, but if the concentration of NaCl can be more precisely controlled, it is thought that the separation of COS having a molecular size of (GlcN)₇ or larger may be possible. Prior to separation and purification of single molecules, it is also necessary to improve methodological approaches that can efficiently produce degradation products sufficiently containing molecules of a certain size. Therefore, the results of this study emphasize once again the importance of the production method of COS of various sizes according to enzyme treatment, and at the same time suggest the need to establish a method for purifying COS of a specific molecular size for pharmaceutical applications.

Detection of Campylobacter jejuni and Salmonella typhimurium in chicken using PCR for virulence factor hipO and invA genes (Saudi Arabia).

Campylobacter jejuni and Salmonella typhimurium are the leading causes of bacterial food contamination in chicken carcasses. Contamination is particularly associated with the slaughtering process. The present study isolated C. jejuni and S. typhimurim from fifty chicken carcass samples, all of which were acquired from different companies in Riyadh, Saudi Arabia. The identification of C. jejuni was performed phenotypically by using a hippurate test and genetically using a polymerase chain reaction with primers for 16S rRNA and hippurate hydrolase (hipO gene). For the dentification of S. typhimurim, a serological Widal test was carried out using serum anti-S. typhimurium antibodies. Strains were genetically detected using invA gene primers. The positive isolates for C. jejuni showed a specific molecular size of 1448 bp for 16S rRNA and 1148 bp for hipO genes. However, the positive isolates of the invA gene exhibited a specific molecular size at 244 bp using polymerase chain reaction (PCR). Comparing sequencing was performed with respect to the invA gene and the BLAST nucleotide isolates that were identified as Salmonella enterica subsp. enterica serovar typhimurium strain ST45, thereby producing a similarity of 100%. The testing identified C. jejuni for hippuricase, GenBank: Z36940.1. While many isolates of Salmonella spp. that contained the invA gene were not necessarily identified as S. typhimurim, the limiting factor for the Widal test used antiS. typhimurum antibodies. The multidrug resistance (MDR) of C. jejuni isolates in chickens was compared with the standard C. jejuni strain ATCC 22931. Similarly, S. typhimurium isolates were compared with the standard S. typhimurium strain ATCC 14028.

A Purified Glucomannan Oligosaccharide from Amorphophallus konjac Improves Colonic Mucosal Barrier Function via Enhancing Butyrate Production and Histone Protein H3 and H4 Acetylation.

A structurally defined konjac glucomannan oligosaccharide (KGMOS) with a relatively high molecular weight and narrow molecular weight distribution (molecular weight ranging from 3000 to 4000 Da, degree of polymerization (dp) 8-11) was prepared from native konjac glucomannan (KGM), and the beneficial effects and molecular mechanisms of KGMOS on colonic functions were investigated in C57BL/6 mice. The results are the first to reveal that KGMOS regulated intestinal microflora composition to facilitate the production of colonic butyrate. Elevated butyrate production further increased the acetylation of histone proteins H3 and H4 and thus enhanced the transcription of the major colonic mucin gene Muc2 and the secretion of mucin elements, which represents a new molecular mechanism of KGM oligosaccharide consumption. The findings indicate that KGM oligosaccharides with specific molecular sizes have highly desirable functional properties and potentially could improve gut health by promoting the barrier function of the colonic mucosa.

Fabrication of interlayer-free silica-based membranes – effect of low calcination temperature using an organo-catalyst

Communities in some wetland areas in South Kalimantan, Indonesia, find it difficult to access clean water because surface water is contaminated by saltwater intrusion, especially during the rainy season. Membrane technology provides a solution to this problem by removing the salt contained in the raw water. The membranes used are generally composed of a porous material – thin-film – medium that functions to separate particles with a specific molecular size from the solution. The purpose of this study is to synthesise and fabricate silica-based membranes using an organo-catalyst calcined at a low calcination temperature (200°C and 250°C) through rapid thermal processing. In order to fabricate these membranes, the sol–gel method was applied, using tetraethyl orthosilicate as the silica precursor and citric acid as an organo-catalyst (refluxed at 0°C and 50°C). The results show that all membranes prepared with the organo-catalyst produce mesoporous structures. The highest surface area and pore size achieved were 354 m2/g and 0.215 cm3/g, respectively, refluxed at 0°C. Furthermore, silica-based membranes calcined at 250°C showed lower silanol concentration compared with those calcined at 200°C. In addition, the organo-catalyst (citric acid) acts as a carbon source in the silica matrices and may increase the hydrostability of the silica networks.

Multiplex RT-PCR method for the simultaneous detection of nine grapevine viruses.

Viral diseases are a serious pathological problem for grapevines, and in recent years the need for increasingly specific and rapid diagnostic methods for the selection of propagation materials has grown. Arabis mosaic virus, Grapevine fanleaf virus, Grapevine virus A, Grapevine virus B, Grapevine rupestris stem pitting-associated virus, Grapevine fleck virus, and Grapevine leafroll-associated viruses 1, 2, and 3 are nine of the most widespread viruses that naturally infect grapevines. A multiplex RT-PCR was developed for simultaneous detection of these nine grapevine viruses, in combination with a plant RNA internal control used as an indicator of the effectiveness of the reaction. One to ten fragments specific for the viruses and an internal control were simultaneously amplified from infected samples and identified by their specific molecular sizes in agarose gel. The protocol reported is an update of previously published protocols for RNA extraction and multiplex diagnosis of viruses. After several years of use and hundreds of samples tested, and following validation in several laboratories, this multiplex RT-PCR provides a reliable and rapid method for detecting grapevine viruses from a large number of samples.

Source and Characteristic of Fluorescence Humic Substances in Arsenic Polluted Groundwater of Bangladesh

AbstractFluorescence characteristic and molecular size distribution of groundwater humic substance (HS) from arsenic contaminated (AsCon) and uncontaminated (nAsCon) areas of Bangladesh indicated that source of HS in aquifers are different with specific molecular size distributions. AsCon area showed signature of microbial degraded HS with small molecular size (0.65 KDa) whereas nAsCon area has minor signature of microbial originated HS with relatively high molecular size (1.24 KDa).

Distance control of electromigration-induced silver nanogaps.

Electromigration (EM) in Ag nanocontacts (NCs) was observed in situ on an atomic scale using simultaneous measurements of electrical conductance and mechanical stress. The in situ observations showed that the critical bias voltage of EM was 45 mV. As the bias voltage was increased to 100-200 mV, the NCs broke and gaps with distances of 1.3 +/- 0.8 nm were obtained for NCs having widths smaller than 6 nm. When the bias voltage was further increased to 200-300 mV, the gaps expanded to more than 3 nm, regardless of the NC width. It was found that the nanogap distance could be controlled to fit specific molecular sizes by appropriately selecting the bias voltage and NC width.

Degradation and humification of maize straw in soil microcosms inoculated with simple and complex microbial communities

SummaryMicrobial communities are responsible for soil organic matter cycling and thus for maintaining soil fertility. A typical Orthic Luvisol was freed from organic carbon by thermal destruction at 600°C. Then the degradation and humification of 14C‐labelled maize straw by defined microbial communities was analysed. To study the role of microbial diversity on the humification of plant material, microcosms containing sterilized soil were inoculated with a natural microbial community or with microbial consortia consisting of bacterial and fungal soil isolates. Within 6 weeks, 41 ± 4% of applied 14C‐labelled maize straw was mineralized in the soil microcosms containing complex communities derived from a soil suspension, whilst the most efficient communities composed of soil isolates mineralized less than 35%. The humification products were analysed by solution state 13C‐NMR‐spectroscopy and gel permeation chromatography (GPC). The analyses of humic acids extracts by solution state 13C‐NMR‐spectroscopy revealed no difference in the development of typical chemical functional groups for humic substances during incubation. However, the increase in specific molecular size fractions of the extracted humic acids occurred only after inoculation with complex communities, but not with defined isolates. While it seems to be true that redundancy in soil microbial communities contributes to the resilience of soils, specific soil functions may no longer be performed if a microbial community is harshly affected in its diversity or growth conditions.

Simultaneous Detection of Nine Grapevine Viruses by Multiplex Reverse Transcription-Polymerase Chain Reaction with Coamplification of a Plant RNA as Internal Control

ABSTRACT A multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was developed for simultaneous detection of nine grapevine viruses: Arabis mosaic virus, Grapevine fanleaf virus, Grapevine virus A, Grapevine virus B, Rupestris stem pitting-associated virus, Grapevine fleck virus, Grapevine leafroll-associated virus-1, -2, and -3, in combination with a plant RNA internal control used as an indicator of the effectiveness of RNA extraction and RT-PCR. Primers were designed from conserved regions of each virus and their specificity was confirmed by sequencing PCR products. Two plant total RNA extraction methods (silica capture and modified RNeasy method) and two RT-PCR systems (onestep and two-step) were evaluated to develop a reliable protocol for mRT-PCR. One to nine fragments specific for the viruses were simultaneously amplified from infected samples and identified by their specific molecular sizes in agarose gel electrophoresis. In the two-step mRT-PCR, the detection limits were 10(-3) or 10(-4) extract dilutions, depending on the virus. Leaves, phloem from dormant cuttings, and in vitro plantlets from 103 naturally infected and healthy grapevines were analyzed. The mRT-PCR provided a reliable and rapid method for detecting grapevine viruses from a large number of samples.

Sorption behavior of plutonium(IV) onto soils in the presence of humic acid

Influence of humic acid on the sorption of 238Pu onto geologic media using a coastal sand, which does not sorb humic acid, and an ando soil, which sorbs humic acid very well, was examined with respect to molecular sizes of humic acid. Apparent sorption affinity of 238Pu for the coastal sand decreased with increasing humic acid concentration. As to the ando soil, the sorption affinity of 238Pu became largest in the presence of several mg/dm3 of humic acid. The humic acid fraction smaller than 100,000 daltons relates to the complexation with 238Pu and the sorption onto the soils, particularly the humic acid fraction of 30,000-100,000 daltons was dominant. Most 238Pu formed humic compounds even in a low humic acid concentration region; resulting plutonium humates are considered to control the apparent sorption affinity of ~38Pu. It is concluded from above results that a specific molecular size of humic acid controls the complexation ability and the apparent sorption affinity of 238Pu.

Simultaneous detection of six citrus viroids and Apple stem grooving virus from citrus plants by multiplex reverse transcription polymerase chain reaction

We developed a multiplex reverse transcription polymerase chain reaction (RT-PCR) to detect six citrus viroids: Citrus exocortis viroid (CEVd), Citrus bent leaf viroid (CBLVd), Hop stunt viroid (HSVd), Citrus viroid III (CVd-III), Citrus viroid IV (CVd-IV) and Citrus viroid OS (CVd-OS) and Apple stem grooving virus (ASGV, synonym: Citrus tatter leaf virus (CTLV)) from citrus plants. The multiplex RT-PCR was also designed to distinguish CVd-I-LSS (a distinct variant of CBLVd) from CBLVd. By the multiplex RT-PCR, one to eight fragments specific to the pathogens were simultaneously amplified from one sample and identified by their specific molecular sizes in 6% polyacrylamide gel electrophoresis. The results of the multiplex RT-PCR were consistent with those of other diagnoses, such as uniplex RT-PCR, to detect each of the pathogens. The multiplex RT-PCR provides a simple and rapid method for detecting various viroids and ASGV in citrus plants, which will help diagnose many citrus plants at a time.

Polymerization kinetics of wheat gluten upon thermosetting. A mechanistic model.

Size exclusion high-performance liquid chromatography analysis was carried out on wheat gluten-glycerol blends subjected to different heat treatments. The elution profiles were analyzed in order to follow the solubility loss of protein fractions with specific molecular size. Owing to the known biochemical changes involved during the heat denaturation of gluten, a mechanistic mathematical model was developed, which divided the protein denaturation into two distinct reaction steps: (i) reversible change in protein conformation and (ii) protein precipitation through disulfide bonding between initially SDS-soluble and SDS-insoluble reaction partners. Activation energies of gluten unfolding, refolding, and precipitation were calculated with the Arrhenius law to 53.9 kJ x mol(-1), 29.5 kJ x mol(-1), and 172 kJ x mol(-1), respectively. The rate of protein solubility loss decreased as the cross-linking reaction proceeded, which may be attributed to the formation of a three-dimensional network progressively hindering the reaction. The enhanced susceptibility to aggregation of large molecules was assigned to a risen reaction probability due to their higher number of cysteine residues and to the increased percentage of unfolded and thereby activated proteins as complete protein refolding seemed to be an anticooperative process.

Analysis of lupin seed protein digestibility using gel electrophoresis and immunoblots.

Proteins from the seeds of 12 cultivars of three lupin species were analyzed by gel electrophoresis. Similarities between cultivars of the same species were noted. Antibodies raised against the three major globular proteins, conglutin alpha, beta, and gamma, of Lupinus albus cv. Ultra were used to probe immunoblots of crude extracts. The immunoblots revealed variations between cultivars not previously resolved and identified which protein-subunits were derived from which conglutin. In vitro digestibility studies were done on four of the lupin cultivars. During the digestion of these cultivars, the large protein units were shown to be degraded to smaller intermediates with specific molecular sizes. Some of the intermediate protein subunits were identified as being derived from conglutin beta. The digestibility of the four cultivars, based on the amount of identifiable protein in the ruminal fluid digest at 9 and 24 h, showed Ultra > Primorski > Juno > Danja. From this study a novel system of analyzing protein digestibility was devised.

Reminiscences and discoveries: Dorothy Hodgkin and the Indian connection

Dorothy Hodgkin - as crystallographer, scientist and human being - far surpasses most, and so it is not easy to write about her many-splendoured personality. Instead, my aim here will he to discuss her influence on the growth of X-ray crystallography in India, directly through those who worked with her and indirectly by her travelling all over this country. In such an account, one must be pardoned for some personal element creeping in. In the twenties, India had developed a fairly strong tradition in X-ray physics. The six-week visit of C.V. Raman to Europe in 1921 greatly changed his research interests. On seeing the blue of the Mediterranean he started his researches on the scattering of light in liquids which finally culminated in the discovery of what is now called the Raman Effect. His encounter with Sir William Bragg and his work on naphthalene structure started three lines of research in India. First, Raman fabricated an X-ray tube and was amongst the earliest to use X-ray diffraction as a structural tool to study liquids. He showed that while in large-angle scattering the haloes reflected specific molecular sizes and packing shapes, small-angle scattering was directly related to the statistical fluctuation of density in a liquid. Second, Raman knew that Bragg’s first structure of naphthalene was not consistent with its birefringence, while the second one was. With this as cue he and his school launched extensive studies on the optical and magnetic anisotropy of organic crystals to get vital information on the arrangements of molecules in the crystalline state. Third, one of his students, Kedareshwar Bannerjee, was amongst the earliest to probe into the problem of phase determination by direct methods and for this he used Bragg’s data on naphthalene. Unfortunately, in spite of this early lead, it was not until 1951 that the first crystal structure was solved in India using Fourier methods by Gopinath Kartha. The Indian Institute of Science (IISc) had great hopes of starting a powerful school of X-ray crystallography when G.N. Ramachandran came back from Cambridge. But he went over to Madras, and there he established one of the most renowned Schools of Biophysics. With Gopinath Kartha he solved the structure of collagen.

The heterogeneity of the protein content of liver and muscle glycogens

The portions of both liver and muscle glycogen that have a high protein content have been investigated. In liver the high molecular weight protions of glycogen may be rendered insoluble by treatment with trichloroacetic acid. This shows that reported desmo- (or insoluble) glycogen is an artefact of the extraction process and therefore of no physiological significance. In contrast, muscle glycogen isolubility is not associated with any specific molecular size range. Insolubility of muscle glycogen is shown to be related to partial degradation of the polysaccharide and to the high protein content remaining after the gentle extraction procedure. Since the molecular weight profile is unaltered by the removal of the insoluble glycogen it does not interfere with the interpretation of metabolic studies.