Displaying publications 1 - 20 of 523 in total

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  1. Mahlangu JN, Weldingh KN, Lentz SR, Kaicker S, Karim FA, Matsushita T, et al.
    J Thromb Haemost, 2015 Nov;13(11):1989-98.
    PMID: 26362483 DOI: 10.1111/jth.13141
    BACKGROUND: Vatreptacog alfa, a recombinant human factor VIIa (rFVIIa) analog developed to improve the treatment of bleeds in hemophilia patients with inhibitors, differs from native FVIIa by three amino acid substitutions. In a randomized, double-blind, crossover, confirmatory phase III trial (adept(™) 2), 8/72 (11%) hemophilia A or B patients with inhibitors treated for acute bleeds developed anti-drug antibodies (ADAs) to vatreptacog alfa.

    OBJECTIVES: To characterize the formation of anti-vatreptacog alfa ADAs in hemophilia patients with inhibitors.

    METHODS/PATIENTS: This was a post hoc analysis of adept(™) 2. Immunoglobulin isotype determination, specificity analysis of rFVIIa cross-reactive antibodies, epitope mapping of rFVIIa single mutant analogs and pharmacokinetic (PK) profiling were performed to characterize the ADAs.

    RESULTS: Immunoglobulin isotyping indicated that the ADAs were of the immunoglobulin G subtype. In epitope mapping, none of the rFVIIa single mutant analogs (V158D, E296V or M298Q) contained the complete antibody epitope, confirming that the antibodies were specific for vatreptacog alfa. In two patients, for whom PK profiling was performed both before and after the development of ADAs, vatreptacog alfa showed a prolonged elimination phase following ADA development. During the follow-up evaluation, the rFVIIa cross-reactivity disappeared after the last vatreptacog alfa exposure, despite continued exposure to rFVIIa as part of standard care.

    CONCLUSIONS: Results from the vatreptacog alfa phase III trial demonstrate that the specific changes made, albeit relatively small, to the FVIIa molecule alter its clinical immunogenicity.

    Matched MeSH terms: Amino Acid Sequence
  2. Hussain A, Khan MI, Albaqami M, Mahpara S, Noorka IR, Ahmed MAA, et al.
    Int J Mol Sci, 2021 Nov 08;22(21).
    PMID: 34769521 DOI: 10.3390/ijms222112091
    The WRKY transcription factors (TFs) network is composed of WRKY TFs' subset, which performs a critical role in immunity regulation of plants. However, functions of WRKY TFs' network remain unclear, particularly in non-model plants such as pepper (Capsicum annuum L.). This study functionally characterized CaWRKY30-a member of group III Pepper WRKY protein-for immunity of pepper against Ralstonia solanacearum infection. The CaWRKY30 was detected in nucleus, and its transcriptional expression levels were significantly upregulated by R. solanacearum inoculation (RSI), and foliar application ethylene (ET), abscisic acid (ABA), and salicylic acid (SA). Virus induced gene silencing (VIGS) of CaWRKY30 amplified pepper's vulnerability to RSI. Additionally, the silencing of CaWRKY30 by VIGS compromised HR-like cell death triggered by RSI and downregulated defense-associated marker genes, like CaPR1, CaNPR1, CaDEF1, CaABR1, CaHIR1, and CaWRKY40. Conversely, transient over-expression of CaWRKY30 in pepper leaves instigated HR-like cell death and upregulated defense-related maker genes. Furthermore, transient over-expression of CaWRKY30 upregulated transcriptional levels of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40. On the other hand, transient over-expression of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40 upregulated transcriptional expression levels of CaWRKY30. The results recommend that newly characterized CaWRKY30 positively regulates pepper's immunity against Ralstonia attack, which is governed by synergistically mediated signaling by phytohormones like ET, ABA, and SA, and transcriptionally assimilating into WRKY TFs networks, consisting of CaWRKY6, CaWRKY22, CaWRKY27, and CaWRKY40. Collectively, our data will facilitate to explicate the underlying mechanism of crosstalk between pepper's immunity and response to RSI.
    Matched MeSH terms: Amino Acid Sequence
  3. Salahshourifar I, Halim AS, Wan Sulaiman WA, Zilfalil BA
    J Hum Genet, 2011 Nov;56(11):755-8.
    PMID: 21866112 DOI: 10.1038/jhg.2011.95
    Oral clefts are clinically and genetically heterogeneous disorders that are influenced by both genetic and environmental factors. The present family-based association study investigated the role of the MSX1 and TGFB3 genes in the etiology of non-syndromic oral cleft in a Malay population. No transmission distortion was found in the transmission disequilibrium analysis for either MSX1-CA or TGFB3-CA intragenic markers, whereas TGFB3-CA exhibited a trend to excess maternal transmission. In sequencing the MSX1 coding regions in 124 patients with oral cleft, five variants were found, including three known variants (A34G, G110G and P147Q) and two novel variants (M37L and G267A). The P147Q and M37L variants were not observed in 200 control chromosomes, whereas G267A was found in one control sample, indicating a very rare polymorphic variant. Furthermore, the G110G variant displayed a significant association between patients with non-syndromic cleft lip, with or without cleft palate, and normal controls (P=0.001, odds ratio=2.241, 95% confidence interval, 1.357-3.700). Therefore, these genetic variants may contribute, along with other genetic and environmental factors, to this condition.
    Matched MeSH terms: Amino Acid Sequence
  4. Zhang T, Wu Q, Zhang Z
    Curr Biol, 2020 04 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
  5. Chen L, Yao XJ, Xu SJ, Yang H, Wu CL, Lu J, et al.
    Arch Virol, 2018 Nov 29.
    PMID: 30498962 DOI: 10.1007/s00705-018-4112-3
    Coxsackievirus A16 (CV-A16) of the genotypes B1a and B1b have co-circulated in mainland China in the past decades. From 2013 to 2017, a total of 3,008 specimens from 3,008 patients with mild hand, foot, and mouth disease were collected in the present study. Viral RNA was tested for CV-A16 by a real-time RT-PCR method, and complete VP1 sequences and full-length genome sequences of CV-A16 strains from this study were determined by RT-PCR and sequencing. Sequences were analyzed using a series of bioinformatics programs. The detection rate for CV-A16 was 4.1%, 25.9%, 10.6%, 28.1% and 12.9% in 2013, 2014, 2015, 2016 and 2017, respectively. Overall, the detection rate for CV-A16 was 16.5% (497/3008) in this 5-year period in Shenzhen, China. One hundred forty-two (142/155, 91.6%) of the 155 genotype B1 strains in the study belonged to subgenotype B1b, and 13 (13/155, 8.4%) strains belonged to subgenotype B1a. Two strains (CVA16/Shenzhen174/CHN/2017 and CVA16/Shenzhen189/CHN/2017) could not be assigned to a known genotype. Phylogenetic analysis of these two strains and other Chinese CV-A16 strains indicated that these two CV-A16 strains clustered independently in a novel clade whose members differed by 8.4%-11.8%, 8.4%-12.1%, and 14.6%-14.8% in their nucleotide sequences from those of Chinese B1a, B1b, and genotype D strains, respectively. Phylogenetic analysis of global CV-A16 strains further indicated that the two novel CV-A16 strains from this study grouped in a previously uncharacterized clade, which was designated as the subgenogroup B3 in present study. Meanwhile, phylogenetic reconstruction revealed two other new genotypes, B1d and B4, which included a Malaysian strain and two American strains, respectively. The complete genome sequences of the two novel CV-A16 strains showed the highest nucleotide sequence identity of 92.3% to the Malaysian strain PM-15765-00 from 2000. Comparative analysis of amino acid sequences of the two novel CV-A16 strains and their relatives suggested that variations in the nonstructural proteins may play an important role in the evolution of modern CV-A16.
    Matched MeSH terms: Amino Acid Sequence
  6. Ee SF, Mohamed-Hussein ZA, Othman R, Shaharuddin NA, Ismail I, Zainal Z
    ScientificWorldJournal, 2014;2014:840592.
    PMID: 24678279 DOI: 10.1155/2014/840592
    Polygonum minus is an aromatic plant, which contains high abundance of terpenoids, especially the sesquiterpenes C15H24. Sesquiterpenes were believed to contribute to the many useful biological properties in plants. This study aimed to functionally characterize a full length sesquiterpene synthase gene from P. minus. P. minus sesquiterpene synthase (PmSTS) has a complete open reading frame (ORF) of 1689 base pairs encoding a 562 amino acid protein. Similar to other sesquiterpene synthases, PmSTS has two large domains: the N-terminal domain and the C-terminal metal-binding domain. It also consists of three conserved motifs: the DDXXD, NSE/DTE, and RXR. A three-dimensional protein model for PmSTS built clearly distinguished the two main domains, where conserved motifs were highlighted. We also constructed a phylogenetic tree, which showed that PmSTS belongs to the angiosperm sesquiterpene synthase subfamily Tps-a. To examine the function of PmSTS, we expressed this gene in Arabidopsis thaliana. Two transgenic lines, designated as OE3 and OE7, were further characterized, both molecularly and functionally. The transgenic plants demonstrated smaller basal rosette leaves, shorter and fewer flowering stems, and fewer seeds compared to wild type plants. Gas chromatography-mass spectrometry analysis of the transgenic plants showed that PmSTS was responsible for the production of β -sesquiphellandrene.
    Matched MeSH terms: Amino Acid Sequence
  7. Kusumaningtyas E, Tan WS, Zamrod Z, Eshaghi M, Yusoff K
    Arch Virol, 2004 Sep;149(9):1859-65.
    PMID: 15593426
    Nucleotide sequence comparison of the L gene of the Malaysian neurotropic-viscerotropic velogenic NDV strain AF2240 with other NDV strains revealed a single nucleotide insertion at position 3870. This mutation is compensated by a nucleotide deletion downstream at position 3958 which results in two forms of the L proteins containing a 30-amino acid substitution in Domain V. This compensatory mutation does not correlate with the pathogenicity of the viral strains but it may affect the viral replication as Domain V is believed to play an important role in the replication of paramyxoviruses.
    Matched MeSH terms: Amino Acid Sequence
  8. Eshaghi M, Tan WS, Yusoff K
    J Med Virol, 2005 Jan;75(1):147-52.
    PMID: 15543570
    A random peptide library of heptamers displayed on the surface of M13 bacteriophage was used to identify specific epitopes of antibodies in pooled sera of swine naturally infected by Nipah virus. The selected heptapeptides were aligned with protein sequences of Nipah virus and several putative epitopes were identified within the nucleocapsid protein. A total of 41 of 60 (68%) selected phage clones had inserts resembling a region with the sequence SNRTQGE, located at the C-terminal end (amino acids 503-509) of the nucleocapsid protein. The binding of antibodies in the swine and human antisera to the phage clone was inhibited by a synthetic peptide corresponding to this region. Epitopes identified by phage display are consistent with the predicted antigenic sites for the Nipah virus nucleocapsid protein. The selected phage clone used as a coating antigen discriminated swine and human Nipah virus sera-positive from sera-negative samples exhibiting characteristics, which might be attractive for diagnostic tests.
    Matched MeSH terms: Amino Acid Sequence
  9. Eshaghi M, Ali AM, Jamal F, Yusoff K
    J. Biochem. Mol. Biol. Biophys., 2002 Feb;6(1):23-8.
    PMID: 12186779
    Streptococcus pyogenes ST4547 is an opacity factor negative strain, which has been recently reported as a new emm type from Malaysia. Nucleotide sequencing of the mga regulon of this strain showed the existence of two emm-like genes. The emm gene located upstream of the scpA gene comprises 1305 nucleotides encoding the putative precursor M protein of 435 amino acids in length with an M(r) of 49 kDa. or a predicted mature protein of 394 amino acids with an M(r) of 44.8 kDa. Another gene mrpST4547 was located upstream of the emm gene and downstream of the mga gene. The sequence of this mrp gene comprises 1167 nucleotides encoding a predicted protein of 388 amino acids in length with an M(r) of 42.2 kDa. or a predicted mature protein of 347 amino acids with an M(r) of 37.9 kDa. The mga regulon of strain ST4547 has a mosaic structure comprising segments, which originated from different OF positive and OF negative strains. The sequences flanking the hyper-variable and C repeats of the emmST4547 gene showed high similarity to corresponding regions in the mga regulon of OF positive strains notably M15, M4, M22 and M50. In contrast, the sequence within the hyper-variable and C repeat regions of the emmST4547 gene revealed high similarity to equivalent regions in the OF negative strains. These data indicates that horizontal transfer of emm-like gene could have occurred between OF positive and OF negative strains resulting in architectural divergence in the mga regulon.
    Matched MeSH terms: Amino Acid Sequence
  10. Kho CL, Tan WS, Yusoff K
    J. Biochem. Mol. Biol. Biophys., 2002 Apr;6(2):117-21.
    PMID: 12186767
    The phosphoprotein (P) gene of a heat stable Newcastle disease virus (NDV) was cloned, sequenced and expressed in Escherichia coli. SDS-PAGE analysis of the recombinant P protein (395 amino acids) and a C-terminal extension derivative (424 amino acids), gave rise to two distinct protein bands with molecular masses of approximately 53-55 and 56-58 kDa, respectively, which are approximately 26-30% heavier than those calculated from the deduced amino acid sequences. The differences in molecular mass on SDS-PAGE are thought to be attributed to the acidic nature of the P protein (pI=6.27) and also the different degrees of phosphorylation in the prokaryotic cell. Amino acid sequence comparison of the P protein among the published NDV strains showed that they were highly conserved particularly at the putative phosphorylation sites.
    Matched MeSH terms: Amino Acid Sequence
  11. Tan WS, Lau CH, Ng BK, Ibrahim AL, Yusoff K
    DNA Seq., 1995;6(1):47-50.
    PMID: 8746461
    The nucleotide sequence of the haemagglutinin-neuraminidase (HN) gene of Newcastle disease virus (NDV) viscerotropic-velogenic strain AF2240 was determined by direct RNA sequencing and by sequencing RT-PCR products. It encodes a single open reading frame of 581 amino acids with a calculated Mr of 63.8 kDa. The predicted sequence contains five asparagine glycosylation sites. Comparison of the AF2240 HN protein sequence with 13 other previously published sequences showed 88% homology. This HN protein is unique because it lacked the Arg 403 residue which is present in all of the other strains and cannot be grouped under the proposed three size classes of HN proteins in NDV.
    Matched MeSH terms: Amino Acid Sequence
  12. Raha AR, Hooi WY, Mariana NS, Radu S, Varma NR, Yusoff K
    Plasmid, 2006 Jul;56(1):53-61.
    PMID: 16675013
    A small plasmid designated pAR141 was isolated from Lactococcus lactis subsp. lactis M14 and its complete 1,594 base pair nucleotide sequence was determined. Analysis of the sequence indicated that this plasmid does not carry any industrially important determinants besides the elements involved in plasmid replication and control. The transcriptional repressor CopG and replication initiation protein RepB appeared as a single operon. A small countertranscribed RNA (ctRNA) coding region was found between the copG and repB genes. The double strand origin (dso) and single strand origin (sso) of rolling circle replicating (RCR) plasmids were also identified in pAR141, suggesting that this plasmid replicates by rolling circle (RC) mode. This observation was supported by S1 nuclease and Southern hybridization analyses.
    Matched MeSH terms: Amino Acid Sequence
  13. Ab Aziz NA, Salim N, Zarei M, Saari N, Yusoff FM
    Prep Biochem Biotechnol, 2021;51(1):44-53.
    PMID: 32701046 DOI: 10.1080/10826068.2020.1789991
    The study was conducted to determine anti-tyrosinase and antioxidant activities of the extracted collagen hydrolysate (CH) derived from Malaysian jellyfish, Rhopilema hispidum. Collagen was extracted using 1:1 (w:v) 0.1 M NaOH solution at temperature 25 °C for 48 hr followed by treatment of 1:2 (w:v) distilled water for another 24 hr and freeze-dried. The extracted collagen was hydrolyzed using papain at optimum temperature, pH and enzyme/substrate ratio [E/S] of 60 °C, 7.0 and 1:50, respectively. CH was found to exhibit tyrosinase inhibitory activity, DPPH radical scavenging and metal ion-chelating assays up to 64, 28, and 83%, respectively, after 8 hr of hydrolysis process. The molecular weight of CH was found <10 kDa consisting of mainly Gly (19.219%), Glu (10.428%), and Arg (8.848%). The UV-visible spectrum analysis showed a major and minor peak at 218 and 276 nm, accordingly. The FTIR spectroscopy confirmed the amide groups in CH. The SEM images demonstrated spongy and porous structure of CH. In the cytotoxicity study, CH has no cytotoxicity against mouse embryonic 3T3 fibroblast cell line with IC50 value >500 µg/ml. Results revealed that the CH generated from this study has a potential to be developed as active ingredient in cosmeceutical application.
    Matched MeSH terms: Amino Acid Sequence
  14. Rothan HA, Mohamed Z, Suhaeb AM, Rahman NA, Yusof R
    OMICS, 2013 Nov;17(11):560-7.
    PMID: 24044366 DOI: 10.1089/omi.2013.0056
    Dengue virus infects millions of people worldwide, and there is no vaccine or anti-dengue therapeutic available. Antimicrobial peptides have been shown to possess effective antiviral activity against various viruses. One of the main limitations of developing these peptides as potent antiviral drugs is the high cost of production. In this study, high yield production of biologically active plectasin peptide was inexpensively achieved by producing tandem plectasin peptides as inclusion bodies in E. coli. Antiviral activity of the recombinant peptide towards dengue serotype-2 NS2B-NS3 protease (DENV2 NS2B-NS3pro) was assessed as a target to inhibit dengue virus replication in Vero cells. Single units of recombinant plectasin were collected after applying consecutive steps of refolding, cleaving by Factor Xa, and nickel column purification to obtain recombinant proteins of high purity. The maximal nontoxic dose (MNTD) of the recombinant peptide against Vero cells was 20 μM (100 μg/mL). The reaction velocity of DENV2 NS2B-NS3pro decreased significantly after increasing concentrations of recombinant plectasin were applied to the reaction mixture. Plectasin peptide noncompetitively inhibited DENV2 NS2B-NS3pro at Ki value of 5.03 ± 0.98 μM. The percentage of viral inhibition was more than 80% at the MNTD value of plectasin. In this study, biologically active recombinant plectasin which was able to inhibit dengue protease and viral replication in Vero cells was successfully produced in E. coli in a time- and cost- effective method. These findings are potentially important in the development of potent therapeutics against dengue infection.
    Matched MeSH terms: Amino Acid Sequence
  15. Rothan HA, Ambikabothy J, Abdulrahman AY, Bahrani H, Golpich M, Amini E, et al.
    PLoS One, 2015;10(9):e0139248.
    PMID: 26418816 DOI: 10.1371/journal.pone.0139248
    The production of short anticancer peptides in recombinant form is an alternative method for costly chemical manufacturing. However, the limitations of host toxicity, bioactivity and column purification have impaired production in mass quantities. In this study, short cationic peptides were produced in aggregated inclusion bodies by double fusion with a central protein that has anti-cancer activity. The anticancer peptides Tachiplicin I (TACH) and Latarcin 1 (LATA) were fused with the N- and C-terminus of the MAP30 protein, respectively. We successfully produced the recombinant TACH-MAP30-LATA protein and MAP30 alone in E. coli that represented 59% and 68% of the inclusion bodies. The purified form of the inclusion bodies was prepared by eliminating host cell proteins through multiple washing steps and semi-solubilization in alkaline buffer. The purified active protein was recovered by inclusive solubilization at pH 12.5 in the presence of 2 M urea and refolded in alkaline buffer containing oxides and reduced glutathione. The peptide-fusion protein showed lower CC50 values against cancer cells (HepG2, 0.35±0.1 μM and MCF-7, 0.58±0.1 μM) compared with normal cells (WRL68, 1.83±0.2 μM and ARPE19, 2.5±0.1 μM) with outstanding activity compared with its individual components. The presence of the short peptides facilitated the entry of the peptide fusion protein into cancer cells (1.8 to 2.2-fold) compared with MAP30 alone through direct interaction with the cell membrane. The cancer chemotherapy agent doxorubicin showed higher efficiency and selectivity against cancer cells in combination with the peptide- fusion protein. This study provides new data on the mass production of short anticancer peptides as inclusion bodies in E. coli by fusion with a central protein that has similar activity. The product was biologically active against cancer cells compared with normal cells and enhanced the activity and selective delivery of an anticancer chemotherapy agent.
    Matched MeSH terms: Amino Acid Sequence
  16. Rothan HA, Bahrani H, Mohamed Z, Abd Rahman N, Yusof R
    PLoS One, 2014;9(4):e94561.
    PMID: 24722532 DOI: 10.1371/journal.pone.0094561
    Dengue virus (DENV) broadly disseminates in tropical and sub-tropical countries and there are no vaccine or anti-dengue drugs available. DENV outbreaks cause serious economic burden due to infection complications that requires special medical care and hospitalization. This study presents a new strategy for inexpensive production of anti-DENV peptide-fusion protein to prevent and/or treat DENV infection. Antiviral cationic peptides protegrin-1 (PG1) and plectasin (PLSN) were fused with MAP30 protein to produce recombinant antiviral peptide-fusion protein (PG1-MAP30-PLSN) as inclusion bodies in E. coli. High yield production of PG1-MAP30-PLSN protein was achieved by solubilization of inclusion bodies in alkaline buffer followed by the application of appropriate refolding techniques. Antiviral PG1-MAP30-PLSN protein considerably inhibited DENV protease (NS2B-NS3pro) with half-maximal inhibitory concentration (IC50) 0.5±0.1 μM. The real-time proliferation assay (RTCA) and the end-point proliferation assay (MTT assay) showed that the maximal-nontoxic dose of the peptide-fusion protein against Vero cells is approximately 0.67±0.2 μM. The cell-based assays showed considerable inhibition of the peptide-fusion protein against binding and proliferating stages of DENV2 into the target cells. The peptide-fusion protein protected DENV2-challeged mice with 100% of survival at the dose of 50 mg/kg. In conclusion, producing recombinant antiviral peptide-fusion protein by combining short antiviral peptide with a central protein owning similar activity could be useful to minimize the overall cost of short peptide production and take advantage of its synergistic antiviral activities.
    Matched MeSH terms: Amino Acid Sequence
  17. Tai HF, Foo HL, Abdul Rahim R, Loh TC, Abdullah MP, Yoshinobu K
    Microb Cell Fact, 2015;14:89.
    PMID: 26077560 DOI: 10.1186/s12934-015-0280-y
    Bacteriocin-producing Lactic acid bacteria (LAB) have vast applications in human and animal health, as well as in food industry. The structural, immunity, regulatory, export and modification genes are required for effective bacteriocin biosynthesis. Variations in gene sequence, composition and organisation will affect the antimicrobial spectrum of bacteriocin greatly. Lactobacillus plantarum I-UL4 is a novel multiple bacteriocin producer that harbours both plw and plnEF structural genes simultaneous which has not been reported elsewhere. Therefore, molecular characterisation of bacteriocin genes that harboured in L. plantarum I-UL4 was conducted in this study.
    Matched MeSH terms: Amino Acid Sequence
  18. Seyedi SS, Tan SG, Namasivayam P, Yong CSY
    Sains Malaysiana, 2016;45:717-727.
    The Hibiscus sabdariffa var. UMKL (Roselle) investigated here may potentially be used as an alternative fibre source. To
    the best of our knowledge, there was no study focusing on the genetics underlying the cellulose biosynthesis machinery
    in Roselle thus far. This paper presents the results of the first isolation of the cellulose synthase gene, HsCesA1 from this
    plant, which is fundamental for working towards understanding the functions of CesA genes in the cellulose biosynthesis
    of Roselle. A full-length HsCesA1 cDNA of 3528 bp in length (accession no: KJ608192) encoding a polypeptide of 974
    amino acid was isolated. The full-length HsCesA1 gene of 5489 bp length (accession no: KJ661223) with 11-introns
    and a promoter region of 737 bp was further isolated. Important and conserved characteristics of a CesA protein were
    identified in the HsCesA1 deduced amino acid sequence, which strengthened the prediction that the isolated gene being
    a cellulose synthase belonging to the processive class of the 2-glycosyltransferase family 2A. Relative gene expression
    analysis by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) on young leaf and stem tissues
    found that HsCesA1 had similar levels of gene expression in both tissues. Phylogenetic and Blast analyses also supported
    the prediction that the isolated HsCesA1 may play roles in the cell wall depositions in both leaf and stem tissues.
    Matched MeSH terms: Amino Acid Sequence
  19. Wang LY, Wang YS, Cheng H, Zhang JP, Yeok FS
    Ecotoxicology, 2015 Oct;24(7-8):1705-13.
    PMID: 26044931 DOI: 10.1007/s10646-015-1502-0
    Chitinases in terrestrial plants have been reported these are involved in heavy metal tolerance/detoxification. This is the first attempt to reveal chitinase gene (AcCHI I) and its function on metal detoxification in mangroves Aegiceras corniculatum. RT-PCR and RACE techniques were used to clone AcCHI I, while real-time quantitative PCR was employed to assess AcCHI I mRNA expressions in response to Cadmium (Cd). The deduced AcCHI I protein consists of 316 amino acids, including a signal peptide region, a chitin-binding domain (CBD) and a catalytic domain. Protein homology modeling was performed to identify potential features in AcCHI I. The CBD structure of AcCHI I might be critical for metal tolerance/homeostasis of the plant. Clear tissue-specific differences in AcCHI I expression were detected, with higher transcript levels detected in leaves. Results demonstrated that a short duration of Cd exposure (e.g., 3 days) promoted AcCHI I expression in roots. Upregulated expression was also detected in leaves under 10 mg/kg Cd concentration stress. The present study demonstrates that AcCHI I may play an important role in Cd tolerance/homeostasis in the plant. Further studies of the AcCHI I protein, gene overexpression, the promoter and upstream regulation will be necessary for clarifying the functions of AcCHI I.
    Matched MeSH terms: Amino Acid Sequence
  20. Sunderasan E, Bahari A, Arif SA, Zainal Z, Hamilton RG, Yeang HY
    Clin Exp Allergy, 2005 Nov;35(11):1490-5.
    PMID: 16297147 DOI: 10.1111/j.1365-2222.2005.02371.x
    BACKGROUND:
    Hev b 4 is an allergenic natural rubber latex (NRL) protein complex that is reactive in skin prick tests and in vitro immunoassays. On SDS-polyacrylamide gel electrophoresis (SDS-PAGE), Hev b 4 is discerned predominantly at 53-55 kDa together with a 57 kDa minor component previously identified as a cyanogenic glucosidase. Of the 13 NRL allergens recognized by the International Union of Immunological Societies, the 53-55 kDa Hev b 4 major protein is the only candidate that lacks complete cDNA and protein sequence information.

    OBJECTIVE:
    We sought to clone the transcript encoding the Hev b 4 major protein, and characterize the native protein and its recombinant form in relation to IgE binding.

    METHODS:
    The 5'/3' rapid amplification of cDNA ends method was employed to obtain the complete cDNA of the Hev b 4 major protein. A recombinant form of the protein was over-expressed in Escherichia coli. The native Hev b 4 major protein was deglycosylated by trifluoromethane sulphonic acid. Western immunoblots of the native, deglycosylated and recombinant proteins were performed using both polyclonal antibodies and sera from latex-allergic patients.

    RESULTS:
    The cDNA encoding the Hev b 4 major protein was cloned. Its open reading frame matched lecithinases in the conserved domain database and contained 10 predicted glycosylation sites. Detection of glycans on the Hev b 4 lecithinase homologue confirmed it to be a glycoprotein. The deglycosylated lecithinase homologue was discerned at 40 kDa on SDS-PAGE, this being comparable to the 38.53 kDa mass predicted by its cDNA. Deglycosylation of the lecithinase homologue resulted in the loss of IgE recognition, although reactivity to polyclonal rabbit anti-Hev b 4 was retained. IgE from latex-allergic patients also failed to recognize the non-glycosylated E. coli recombinant lecithinase homologue.

    CONCLUSION:
    The IgE epitopes of the Hev b 4 lecithinase homologue reside mainly in its carbohydrate moiety, which also account for the discrepancy between the observed molecular weight of the protein and the value calculated from its cDNA.
    Matched MeSH terms: Amino Acid Sequence
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