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Sequence analysis and structure prediction of type II Pseudomonas sp. USM 4-55 PHA synthase and an insight into its catalytic mechanism

Wahab HA, Ahmad Khairudin NB, Samian MR, Najimudin N

BMC Struct Biol, 2006;6:23.
PMID: 17076907

Polyhydroxyalkanoates (PHA), are biodegradable polyesters derived from many microorganisms such as the pseudomonads. These polyesters are in great demand especially in the packaging industries, the medical line as well as the paint industries. The enzyme responsible in catalyzing the formation of PHA is PHA synthase. Due to the limited structural information, its functional properties including catalysis are lacking. Therefore, this study seeks to investigate the structural properties as well as its catalytic mechanism by predicting the three-dimensional (3D) model of the Type II Pseudomonas sp. USM 4-55 PHA synthase 1 (PhaC1P.sp USM 4-55).

Matched MeSH terms: Amino Acid Sequence
Fulltext Isolation and expression analysis of novel silicon absorption gene from roots of mangrove (Rhizophora apiculata) via suppression subtractive hybridization

Sahebi M, Hanafi MM, Abdullah SN, Rafii MY, Azizi P, Nejat N, et al.

Biomed Res Int, 2014;2014:971985.
PMID: 24516858 DOI: 10.1155/2014/971985

Silicon (Si) is the second most abundant element in soil after oxygen. It is not an essential element for plant growth and formation but plays an important role in increasing plant tolerance towards different kinds of abiotic and biotic stresses. The molecular mechanism of Si absorption and accumulation may differ between plants, such as monocotyledons and dicotyledons. Silicon absorption and accumulation in mangrove plants are affected indirectly by some proteins rich in serine and proline amino acids. The expression level of the genes responsible for Si absorption varies in different parts of plants. In this study, Si is mainly observed in the epidermal roots' cell walls of mangrove plants compared to other parts. The present work was carried out to discover further information on Si stress responsive genes in Rhizophora apiculata, using the suppression subtractive hybridization technique. To construct the cDNA library, two-month-old seedlings were exposed to 0.5, 1, and 1.5 mM SiO2 for 15 hrs and for 1 to 6 days resulting in a total of 360 high quality ESTs gained. Further examination by RT-PCR and real-time qRT-PCR showed the expression of a candidate gene of serine-rich protein.

Matched MeSH terms: Amino Acid Sequence
Immunoaffinity extraction and tandem mass spectrometric analysis of human chorionic gonadotropin in doping analysis

Gam LH, Tham SY, Latiff A

J Chromatogr B Analyt Technol Biomed Life Sci, 2003 Jul 25;792(2):187-96.
PMID: 12860026

A confirmatory and quantitative HPLC-tandem mass spectrometry (MS-MS) method for human chorionic gonadotropin hormone (hCG) at concentrations as low as 5 IU/l following immunoaffinity extraction of the glycoprotein from urine was developed. The extraction method involved retention of urinary hCG in the immunoaffinity column via specific antigen-antibody interaction. A variety of eluents were then used to quantitatively elute hCG from the immunoaffinity column. Qualitative and quantitative analysis of hCG were undertaken using MS-MS by identifying the amino acid sequence of the marker peptide betaT5 obtained from hCG by tryptic digestion and the peak areas of three product ions b(6)(+), b(9)(+) and y(11)(+), respectively.

Matched MeSH terms: Amino Acid Sequence
An immunogenic epitope of Chlamydia pneumoniae from a random phage display peptide library is reactive with both monoclonal antibody and patients sera

Naidu BR, Ngeow YF, Wang LF, Chan L, Yao ZJ, Pang T

Immunol Lett, 1998 Jun;62(2):111-5.
PMID: 9698107

Random 15-mer peptides displayed on filamentous phages were screened in binding studies using a Chlamydia pneumoniae-specific monoclonal antibody (RR-402) and affinity-purified, polyclonal sera from patients seropositive for C. pneumoniae infections by the microimmunofluorescence (MIF) test. One 15-mer epitope, epitope Cpnl5A (LASLCNPKPSDAPVT) was identified in both the monoclonal and polyclonal screenings, and showed higher ELISA reactivity with C. pneumoniae MIF-positive sera compared to patients with other chlamydial infections, non-chlamydial respiratory infections and normal healthy sera (MIF-negative). Interestingly, epitope Cpnl5A also showed significant (52%) amino acid sequence homology to the 56 kDa type-specific antigen of Rickettsia tsutsugamushi, a protein implicated in the virulence of this organism.

Matched MeSH terms: Amino Acid Sequence
Fulltext Probable Pangolin Origin of SARS-CoV-2 Associated with the COVID-19 Outbreak

Zhang T, Wu Q, Zhang Z

Curr Biol, 2020 Apr 06;30(7):1346-1351.e2.
PMID: 32197085 DOI: 10.1016/j.cub.2020.03.022

An outbreak of coronavirus disease 2019 (COVID-19) caused by the 2019 novel coronavirus (SARS-CoV-2) began in the city of Wuhan in China and has widely spread worldwide. Currently, it is vital to explore potential intermediate hosts of SARS-CoV-2 to control COVID-19 spread. Therefore, we reinvestigated published data from pangolin lung samples from which SARS-CoV-like CoVs were detected by Liu et al. [1]. We found genomic and evolutionary evidence of the occurrence of a SARS-CoV-2-like CoV (named Pangolin-CoV) in dead Malayan pangolins. Pangolin-CoV is 91.02% and 90.55% identical to SARS-CoV-2 and BatCoV RaTG13, respectively, at the whole-genome level. Aside from RaTG13, Pangolin-CoV is the most closely related CoV to SARS-CoV-2. The S1 protein of Pangolin-CoV is much more closely related to SARS-CoV-2 than to RaTG13. Five key amino acid residues involved in the interaction with human ACE2 are completely consistent between Pangolin-CoV and SARS-CoV-2, but four amino acid mutations are present in RaTG13. Both Pangolin-CoV and RaTG13 lost the putative furin recognition sequence motif at S1/S2 cleavage site that can be observed in the SARS-CoV-2. Conclusively, this study suggests that pangolin species are a natural reservoir of SARS-CoV-2-like CoVs.

Matched MeSH terms: Amino Acid Sequence
Immunoinformatic approach for multi-epitope vaccine design against Staphylococcus aureus based on hemolysin proteins

Garrido-Palazuelos LI, Almanza-Orduño AA, Waseem M, Basheer A, Medrano-Félix JA, Mukthar M, et al.

J Mol Graph Model, 2024 Nov;132:108848.
PMID: 39182254 DOI: 10.1016/j.jmgm.2024.108848

Staphylococcus aureus is a common bacterium that causes a variety of infections in humans. This microorganism produces several virulence factors, including hemolysins, which contribute to its disease-causing ability. The treatment of S. aureus infections typically involves the use of antibiotics. However, the emergence of antibiotic-resistant strains has become a major concern. Therefore, vaccination against S. aureus has gained attention as an alternative approach. Vaccination has the advantage of stimulating the immune system to produce specific antibodies that can neutralize bacteria and prevent infection. However, developing an effective vaccine against S. aureus has proven to be challenging. This study aimed to use in silico methods to design a multi-epitope vaccine against S. aureus infection based on hemolysin proteins. The designed vaccine contained four B-cell epitopes, four CTL epitopes, and four HTL epitopes, as well as the ribosomal protein L7/L12 and pan-HLA DR-binding epitope, included as adjuvants. Furthermore, the vaccine was non-allergenic and non-toxic with the potential to stimulate the TLR2-, TLR-4, and TLR-6 receptors. The predicted vaccine exhibited a high degree of antigenicity and stability, suggesting potential for further development as a viable vaccine candidate. The population coverage of the vaccine was 94.4 %, indicating potential widespread protection against S. aureus. Overall, these findings provide valuable insights into the design of an effective multi-epitope vaccine against S. aureus infection and pave the way for future experimental validations.

Matched MeSH terms: Amino Acid Sequence
Identification and Characterization of Entamoeba histolytica Choline Kinase

Chang CH, See Too WC, Lim BH, Few LL

Acta Parasitol, 2024 Mar;69(1):426-438.
PMID: 38172465 DOI: 10.1007/s11686-023-00763-1

PURPOSE: Entamoeba histolytica is one of the death-causing parasites in the world. Study on its lipid composition revealed that it is predominated by phosphatidylcholine and phosphatidylethanolamine. Further study revealed that its phosphorylated metabolites might be produced by the Kennedy pathway. Here, we would like to report on the characterizations of enzymes from this pathway that would provide information for the design of novel inhibitors against these enzymes in future.
METHODOLOGY: E. histolytica HM-1:IMSS genomic DNA was isolated and two putative choline/ethanolamine kinase genes (EhCK1 and EhCK2) were cloned and expressed from Escherichia coli BL21 strain. Enzymatic characterizations were further carried out on the purified enzymes.
RESULTS: EhCK1 and EhCK2 were identified from E. histolytica genome. The deduced amino acid sequences were more identical to its homologues in human (35-48%) than other organisms. The proteins were clustered as ethanolamine kinase in the constructed phylogeny tree. Sequence analysis showed that they possessed all the conserved motifs in choline kinase family: ATP-binding loop, Brenner's phosphotransferase motif, and choline kinase motif. Here, the open reading frames were cloned, expressed, and purified to apparent homogeneity. EhCK1 showed activity with choline but not ethanolamine. The biochemical characterization showed that it had a Vmax of 1.9 ± 0.1 µmol/min/mg. Its Km for choline and ATP was 203 ± 26 µM and 3.1 ± 0.4 mM, respectively. In contrast, EhCK2 enzymatic activity was only detected when Mn2+ was used as the co-factor instead of Mg2+ like other choline/ethanolamine kinases. Highly sensitive and specific antibody against EhCK1 was developed and used to confirm the endogenous EhCK1 expression using immunoblotting.
CONCLUSIONS: With the understanding of EhC/EK importance in phospholipid metabolism and their unique characteristic, EhC/EK could be a potential target for future anti-amoebiasis study.

Matched MeSH terms: Amino Acid Sequence
Occurrences of allergenicity to banana pathogenesis-related-10 (PR10) protein variants

Suhaimi AH, Rajendram A, Khaidizar FD, Mir P, Pulido-Lucas E, Quirce S, et al.

Food Funct, 2024 Nov 25;15(23):11715-11725.
PMID: 39539124 DOI: 10.1039/d4fo03301a

Pathogenesis-related-10 (PR10) proteins play significant roles in plant defence against biotic and abiotic stresses. Recently, two banana PR10 proteins (MaPR10-BeB5 and MaPR10-GNA5) were characterised and shown to exhibit antifungal properties against Aspergillus fumigatus in vitro. In rice, transgenic overexpression of PR10 proteins conferred resistance to pathogen infection and drought tolerance without affecting productivity, highlighting their potential for agricultural applications. However, PR10 proteins also include the Bet v 1-like family of allergens implicated in pollen food allergy syndromes, raising concerns about potential adverse effects on human health. In this study, we evaluated the allergenic potential of the recently isolated banana PR10 proteins. We first predicted the presence of IgE epitopes of the Bet v 1 allergen family in the deduced PR10 peptide sequences in silico. We then predicted the structures of four human IgE scFv protein sequences and three plant PR10 protein sequences. Based on the quality of the predicted structures, one IgE scFv protein structure was selected for docking with the three plant PR10 proteins. We confirmed the docking results with immunoblot analysis performed using recombinant MaPR10-BeB5 and MaPR10-GNA5 proteins against the sera of banana-allergic patients. Our experimental results substantiated the notion that both protein variants are potentially allergenic since these proteins were recognised by 26.6% of banana-allergic patients with broad PR10 protein recognition. We caution that the allergenic potential of MaPR10 proteins should be carefully considered before implementing transgenic overexpression strategies to improve crops, with a suggestion to limit their expression to non-edible plant tissues.

Matched MeSH terms: Amino Acid Sequence
The mannose-binding lectin (MBL) gene cloned from Exopalaemon carinicauda plays a key role in resisting infection by Vibrio parahaemolyticus

Shi T, Gao J, Xu W, Liu X, Yan B, Azra MN, et al.

Comp Biochem Physiol B Biochem Mol Biol, 2024;274:111001.
PMID: 38908544 DOI: 10.1016/j.cbpb.2024.111001

Mannose-binding lectin (MBL) is a vital member of the lectin family, crucial for mediating functions within the complement lectin pathway. In this study, following the cloning of the mannose-binding lectin (MBL) gene in the ridgetail white prawn, Exopalaemon carinicauda, we examined its expression patterns across various tissues and its role in combating challenges posed by Vibrio parahaemolyticus. The results revealed that the MBL gene spans 1342 bp, featuring an open reading frame of 972 bp. It encodes a protein comprising 323 amino acids, with a predicted relative molecular weight of 36 kDa and a theoretical isoelectric point of 6.18. The gene exhibited expression across various tissues including the eyestalk, heart, gill, hepatopancreas, stomach, intestine, ventral nerve cord, muscle, and hemolymph, with the highest expression detected in the hepatopancreas. Upon challenge with V. parahaemolyticus, RT-PCR analysis revealed a trend of MBL expression in hepatopancreatic tissues, characterized by an initial increase followed by a subsequent decrease, peaking at 24 h post-infection. Employing RNA interference to disrupt MBL gene expression resulted in a significant increase in mortality rates among individuals challenged with V. parahaemolyticus. Furthermore, we successfully generated the Pet32a-MBL recombinant protein through the construction of a prokaryotic expression vector for conducting in vitro bacterial inhibition assays, which demonstrated the inhibitory effect of the recombinant protein on V. parahaemolyticus, laying a foundation for further exploration into its immune mechanism in response to V. parahaemolyticus challenges.

Matched MeSH terms: Amino Acid Sequence
Fulltext Virological investigation of genetic variation of enterovirus type 71 in hand, foot and mouth disease

Wang Y, Li Y, Yang Y, Peng C, Fu X, Gu X, et al.

Exp Ther Med, 2020 Jul;20(1):543-549.
PMID: 32537012 DOI: 10.3892/etm.2020.8728

The aim of the present study was to analyze the sequence of the VP1 gene in enterovirus 71 (EV71) isolates and to explore their genetic evolution, so as to provide a scientific basis for the clinical prevention and treatment of hand, foot and mouth disease. The fecal samples of 590 patients with suspected hand, foot and mouth disease treated at Yan'an Hospital (Kunming, China) between January 2015 and December 2016 were collected and EV71 nucleic acid was detected by fluorescence PCR. The viral RNA of EV71-positive samples was extracted, the VP1 gene was amplified by PCR and the products were sequenced. The VP1 gene sequence was analyzed using DNAMAN and MEGA (version 4.0) software and homologous modeling was performed using Pymol software. A total of 50 EV71-positive samples were identified and the detection rate was 8.47% (50/590 cases). All of the 50 EV71 strains were of the C4 subtype. The genetic distance between the strains detected in the present study and EV71 strains detected in Beijing, Anhui and Malaysia was 0.01-0.03, while that between the strains detected in the present study and Australian strains was 2.11. Homologous modeling indicated that the amino acid sequence of the VP1 gene of the detected strains had a H144Y mutation. There was no significant genetic variation in the EV71 strain within the 2-year period. In conclusion, the EV71 strains detected in the present study was similar to that detected in Beijing, Anhui and Malaysia but different to that from Australia. A point mutation was present in the amino acid sequence of the VP1 gene.

Matched MeSH terms: Amino Acid Sequence
Fulltext The structure and dynamics of BmR1 protein from Brugia malayi: in silico approaches

Khor BY, Tye GJ, Lim TS, Noordin R, Choong YS

Int J Mol Sci, 2014 Jun 19;15(6):11082-99.
PMID: 24950179 DOI: 10.3390/ijms150611082

Brugia malayi is a filarial nematode, which causes lymphatic filariasis in humans. In 1995, the disease has been identified by the World Health Organization (WHO) as one of the second leading causes of permanent and long-term disability and thus it is targeted for elimination by year 2020. Therefore, accurate filariasis diagnosis is important for management and elimination programs. A recombinant antigen (BmR1) from the Bm17DIII gene product was used for antibody-based filariasis diagnosis in "Brugia Rapid". However, the structure and dynamics of BmR1 protein is yet to be elucidated. Here we study the three dimensional structure and dynamics of BmR1 protein using comparative modeling, threading and ab initio protein structure prediction. The best predicted structure obtained via an ab initio method (Rosetta) was further refined and minimized. A total of 5 ns molecular dynamics simulation were performed to investigate the packing of the protein. Here we also identified three epitopes as potential antibody binding sites from the molecular dynamics average structure. The structure and epitopes obtained from this study can be used to design a binder specific against BmR1, thus aiding future development of antigen-based filariasis diagnostics to complement the current diagnostics.

Matched MeSH terms: Amino Acid Sequence
Fulltext Structures and binding specificity of galactose- and mannose-binding lectins from champedak: differences from jackfruit lectins

Gabrielsen M, Abdul-Rahman PS, Othman S, Hashim OH, Cogdell RJ

Acta Crystallogr F Struct Biol Commun, 2014 Jun;70(Pt 6):709-16.
PMID: 24915077 DOI: 10.1107/S2053230X14008966

Galactose-binding and mannose-binding lectins from the champedak fruit, which is native to South-east Asia, exhibit useful potential clinical applications. The specificity of the two lectins for their respective ligands allows the detection of potential cancer biomarkers and monitoring of the glycosylated state of proteins in human serum and/or urine. To fully understand and expand the use of these natural proteins, their complete sequences and crystal structures are presented here, together with details of sugar binding.

Matched MeSH terms: Amino Acid Sequence
Fulltext HPV 16E7 and 48E7 proteins use different mechanisms to target p130 to overcome cell cycle block

Nor Rashid N, Yong ZL, Yusof R, Watson RJ

Virol J, 2016 Jan 04;13:2.
PMID: 26728921 DOI: 10.1186/s12985-015-0460-8

Retinoblastoma like protein 2 (RBL2) or p130 is a member of the pocket protein family, which is infrequently mutated in human tumours. Its expression is posttranscriptionally regulated and largely G0 restricted. We have previously shown that E6/E7 oncoproteins encoded by human papillomavirus (HPV) type 16, which is a high-risk type for cervical cancer development, must target p130 to promote the host cell to exit from quiescence (G0) state and enter S phase of the cell cycle. P130 is associated with the DREAM (DP, RB-like, E2F and MuvB) complex in G0/G1, which prevents S phase progression by repressing transcription of E2F-regulated genes. E7 proteins could potentially disrupt the p130-DREAM complex through two known mechanisms: direct interaction with p130 or induction of cyclin dependent kinase 2 (CDK2) phosphorylation by interacting with its inhibitor, p21(CIP1).

Matched MeSH terms: Amino Acid Sequence
Molecular detection and genetic diversity of Babesia gibsoni in dogs in Bangladesh

Terao M, Akter S, Yasin MG, Nakao R, Kato H, Alam MZ, et al.

Infect Genet Evol, 2015 Apr;31:53-60.
PMID: 25620376 DOI: 10.1016/j.meegid.2015.01.011

Babesia gibsoni is a tick-borne hemoprotozoan parasite of dogs that often causes fever and hemolytic illness. Detection of B. gibsoni has been predominantly reported in Asian countries, including Japan, Korea, Taiwan, Malaysia, Bangladesh and India. The present study shows the first molecular characterization of B. gibsoni detected from dogs in Bangladesh. Blood samples were collected on FTA® Elute cards from 50 stray dogs in Mymensingh District in Bangladesh. DNA eluted from the cards was subjected to nested PCR for the 18S rRNA gene of Babesia species. Approximately 800bp PCR products were detected in 15 of 50 dogs (30%). Based on restriction fragment length polymorphism (RFLP) and direct sequencing of the PCR products, all parasite isolates were identified as B. gibsoni. Furthermore, the BgTRAP (B. gibsoni thrombospondin-related adhesive protein) gene fragments were detected in 13 of 15 18S rRNA gene PCR positive blood samples. Phylogenetic analysis of the BgTRAP gene revealed that B. gibsoni parasites in Bangladesh formed a cluster, which was genetically different from other Asian B. gibsoni isolates. In addition, tandem repeat analysis of the BgTRAP gene clearly showed considerable genetic variation among Bangladeshi isolates. These results suggested that B. gibsoni parasites in a different genetic clade are endemic in dogs in Bangladesh. Further studies are required to elucidate the origin, distribution, vector and pathogenesis of B. gibsoni parasites circulating in dogs in Bangladesh.

Matched MeSH terms: Amino Acid Sequence
Molecular characterization, modeling and docking of CYP107CB2 from Bacillus lehensis G1, an alkaliphile

Ang SS, Salleh AB, Chor AL, Normi YM, Tejo BA, Rahman MB

Comput Biol Chem, 2015 Jun;56:19-29.
PMID: 25766878 DOI: 10.1016/j.compbiolchem.2015.02.015

Cytochrome P450s are a superfamily of heme monooxygenases which catalyze a wide range of biochemical reactions. The reactions involve the introduction of an oxygen atom into an inactivated carbon of a compound which is essential to produce an intermediate of a hydroxylated product. The diversity of chemical reactions catalyzed by cytochrome P450s has led to their increased demand in numerous industrial and biotechnology applications. A recent study showed that a gene sequence encoding a CYP was found in the genome of Bacillus lehensis G1, and this gene shared structural similarity with the bacterial vitamin D hydroxylase (Vdh) from Pseudonocardia autotrophica. The objectives of present study was to mine, for a novel CYP from a new isolate B. lehensis G1 alkaliphile and determine the biological properties and functionalities of CYP in this bacterium. Our study employed the usage of computational methods to search for the novel CYP from CYP structural databases to identify the conserved pattern, functional domain and sequence properties of the uncharacterized CYP from B. lehensis G1. A computational homology model of the protein's structure was generated and a docking analysis was performed to provide useful structural knowledge on the enzyme's possible substrate and their interaction. Sequence analysis indicated that the newly identified CYP, termed CYP107CB2, contained the fingerprint heme binding sequence motif FxxGxxxCxG at position 336-345 as well as other highly conserved motifs characteristic of cytochrome P450 proteins. Using docking studies, we identified Ser-79, Leu-81, Val-231, Val-279, Val-383, Ala-232, Thr-236 and Thr-283 as important active site residues capable of stabilizing interactions with several potential substrates, including vitamin D3, 25-hydroxyvitamin D3 and 1α-hydroxyvitamin D3, in which all substrates docked proximally to the enzyme's heme center. Biochemical analysis indicated that CYP107CB2 is a biologically active protein to produce 1α,25-dihydroxyvitamin D3 from 1α-hydroxyvitamin D3. Based on these results, we conclude that the novel CYP107CB2 identified from B. lehensis G1 is a putative vitamin D hydroxylase which is possibly capable of catalyzing the bioconversion of parental vitamin D3 to calcitriol, or related metabolic products.

Matched MeSH terms: Amino Acid Sequence
Fulltext Molecular phylogeny and predicted 3D structure of plant beta-D-N-acetylhexosaminidase

Hossain MA, Roslan HA

ScientificWorldJournal, 2014;2014:186029.
PMID: 25165734 DOI: 10.1155/2014/186029

beta-D-N-Acetylhexosaminidase, a family 20 glycosyl hydrolase, catalyzes the removal of β-1,4-linked N-acetylhexosamine residues from oligosaccharides and their conjugates. We constructed phylogenetic tree of β-hexosaminidases to analyze the evolutionary history and predicted functions of plant hexosaminidases. Phylogenetic analysis reveals the complex history of evolution of plant β-hexosaminidase that can be described by gene duplication events. The 3D structure of tomato β-hexosaminidase (β-Hex-Sl) was predicted by homology modeling using 1now as a template. Structural conformity studies of the best fit model showed that more than 98% of the residues lie inside the favoured and allowed regions where only 0.9% lie in the unfavourable region. Predicted 3D structure contains 531 amino acids residues with glycosyl hydrolase20b domain-I and glycosyl hydrolase20 superfamily domain-II including the (β/α)8 barrel in the central part. The α and β contents of the modeled structure were found to be 33.3% and 12.2%, respectively. Eleven amino acids were found to be involved in ligand-binding site; Asp(330) and Glu(331) could play important roles in enzyme-catalyzed reactions. The predicted model provides a structural framework that can act as a guide to develop a hypothesis for β-Hex-Sl mutagenesis experiments for exploring the functions of this class of enzymes in plant kingdom.

Matched MeSH terms: Amino Acid Sequence
Direct regulation of gonadotropin release by neurokinin B in tilapia (Oreochromis niloticus)

Biran J, Golan M, Mizrahi N, Ogawa S, Parhar IS, Levavi-Sivan B

Endocrinology, 2014 Dec;155(12):4831-42.
PMID: 25211586 DOI: 10.1210/en.2013-2114

Neurokinin B (NKB) was recently identified as a key regulator of reproduction in mammals and fish. Fish were found to possess a specific novel neurokinin termed NKF. To study the role of NKB/NKF in the regulation of fish reproduction and to investigate the role of NKB/NKF and their receptors in the piscine pituitary, we have identified the NKB/tachikinin 3 receptor (tac3r) system in tilapia. Bioinformatics and phylogenetic analyses have demonstrated that the tilapia holds 1 putative tac3 gene and 2 NKB receptor genes (tac3ra and tac3rb) that clustered with other piscine Tac3 and NKB receptor lineages. Furthermore, we found that in African cichlids, NKB peptides differ from other vertebrate NKBs in their C-terminal sequence, possessing isoleucine instead of valine as the X in the NKB FXGLM-NH2-terminal consensus sequence. Signal transduction analysis demonstrated that tilapia NKB (tiNKB), tiNKF, and human NKB activated both CRE-luc and SRE-luc transcriptional activity of both tilapia and human NKB receptors. Two hours after ip injection of tiNKB, the plasma levels of both FSH and LH were increased, whereas tiNKF was more effective in increasing LH levels. However, tiNKB was more effective than tiNKF in increasing both FSH and LH from tilapia pituitary dispersed cells. Using in situ hybridization and fluorescent immunohistochemistry, we have shown that LH cells possess tac3, tac3ra, and tac3rb mRNAs, whereas FSH cells possess mainly tac3rb and tac3ra and tac3 to a much lesser extent. These results suggest that the members of the NKB/tac3r system may serve as paracrine/autocrine regulators of gonadotropin release in fish pituitary.

Matched MeSH terms: Amino Acid Sequence
LPXRFa, the piscine ortholog of GnIH, and LPXRF receptor positively regulate gonadotropin secretion in Tilapia (Oreochromis niloticus)

Biran J, Golan M, Mizrahi N, Ogawa S, Parhar IS, Levavi-Sivan B

Endocrinology, 2014 Nov;155(11):4391-401.
PMID: 25144920 DOI: 10.1210/en.2013-2047

LPXRFamide (LPXRFa) peptides have been characterized for their ability to inhibit gonadotropin (GTH) release in birds and stimulate growth hormone (GH) release in frogs. However, their involvement in regulating the reproductive hypothalamo-pituitary-gonadal axis in mammals and fish is inconclusive. To study the role of LPXRFa peptides in the regulation of GTH secretion, we cloned tilapia LPXRFa and LPXRF receptor (LPXRF-R). Processing of the tilapia preproLPXRFa liberated three mature LPXRFa peptides that varied in size and post-translational modifications. Phylogenetic analysis of LPXRFa and the closely related RFamide peptide PQRFa showed clear clustering of each peptide sequence with its orthologs from various vertebrates. Signal-transduction analysis of the tilapia LPXRF-R in COS-7 cells showed clear stimulation of CRE-dependent luciferase activity, whereas the human NPFFR1 showed suppression of forskolin-induced CRE-dependent activity in this system. Administration of the tilapia pyroglutaminated LPXRFa-2 peptide to primary cell culture of tilapia pituitaries, or to reproductive female tilapia by ip injection, positively regulated both LH and FSH release in vivo and in vitro. Using double-labeled fluorescent in-situ hybridization and immunofluorescence, βLH cells were found to co-express both tilapia lpxrf and tilapia lpxrf-r mRNA, whereas some of the βFSH cells coexpressed only lpxrf-r mRNA. No coexpression of tilapia lpxrf-r was identified in GH-positive cells. These findings suggest that the LPXRFa system is a potent positive regulator of the reproductive neuroendocrine axis of tilapia.

Matched MeSH terms: Amino Acid Sequence
Fulltext Mitochondrial DNA markers reveal high genetic diversity but low genetic differentiation in the black fly Simulium tani Takaoka & Davies along an elevational gradient in Malaysia

Low VL, Adler PH, Takaoka H, Ya'cob Z, Lim PE, Tan TK, et al.

PLoS One, 2014;9(6):e100512.
PMID: 24941043 DOI: 10.1371/journal.pone.0100512

The population genetic structure of Simulium tani was inferred from mitochondria-encoded sequences of cytochrome c oxidase subunits I (COI) and II (COII) along an elevational gradient in Cameron Highlands, Malaysia. A statistical parsimony network of 71 individuals revealed 71 haplotypes in the COI gene and 43 haplotypes in the COII gene; the concatenated sequences of the COI and COII genes revealed 71 haplotypes. High levels of genetic diversity but low levels of genetic differentiation were observed among populations of S. tani at five elevations. The degree of genetic diversity, however, was not in accordance with an altitudinal gradient, and a Mantel test indicated that elevation did not have a limiting effect on gene flow. No ancestral haplotype of S. tani was found among the populations. Pupae with unique structural characters at the highest elevation showed a tendency to form their own haplotype cluster, as revealed by the COII gene. Tajima's D, Fu's Fs, and mismatch distribution tests revealed population expansion of S. tani in Cameron Highlands. A strong correlation was found between nucleotide diversity and the levels of dissolved oxygen in the streams where S. tani was collected.

Matched MeSH terms: Amino Acid Sequence
A human adenovirus species B subtype 21a associated with severe pneumonia

Hage E, Huzly D, Ganzenmueller T, Beck R, Schulz TF, Heim A

J Infect, 2014 Nov;69(5):490-9.
PMID: 24975176 DOI: 10.1016/j.jinf.2014.06.015

Between 2005 and 2013 six severe pneumonia cases (all requiring mechanical ventilation, two fatal outcomes) caused by human adenovirus type 21 (HAdV-B21) were observed in Germany. So far, HAdV-B21 was mainly associated with non-severe upper and lower respiratory tract infections. However, a few highly virulent HAdV types, e.g. HAdV-B14p1, were previously associated with severe, fatal pneumonia. Complete genomic sequences of the German HAdV-B21 pneumonia isolates formed a single phylogenetic cluster with very high sequence identity (≥ 99.897%). Compared to the HAdV-B21 prototype (only 99.319% identity), all isolates had a unique 15 amino acid deletion and a 2 amino acid insertion in the RGD loop of the penton base which may affect binding to the secondary receptor on the host cells. Moreover, a recombinant E4 gene region derived of HAdV-B3 was identified by bootscan analysis. Thus, the highly virulent, pneumotropic HAdV-B21 was denominated as subtype 21a. Surprisingly, there was 99.963% identity with agent Y/SIBU97 (only 13.4 kb available in GenBank of the 35.4 kb genome) which was associated with 10 fatalities due to cardiopulmonary failure in Sarawak, Malaysia, in 1997. In conclusion, a HAdV-B21 subtype (21a) associated with severe pneumonia in Germany was phylogenetically linked to an adenovirus isolated in Malaysia.

Matched MeSH terms: Amino Acid Sequence

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