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1.
Lysozymes function in animal immunity. Three types of lysozyme have been identified in animal kingdom and most lysozymes identified from bivalve molluscs belong to the invertebrate (i) type. In this research, we cloned and sequenced a new i-type lysozyme, named MmeLys, from the Asiatic hard clam Meretrix meretrix. MmeLys cDNA was constituted of 552 bp, with a 441 bp open reading frame encoding a 146 amino acid polypeptide. The encoded polypeptide was predicted to have a 15 amino acid signal peptide, and a 131 amino acid mature protein with a theoretical mass of 14601.44 Da and an isoelectric point (pI) of 7.14. MmeLys amino acid sequence bore 64% identity with the Manila clam (Venerupis philippinarum) i-type lysozyme and was grouped with other veneroid i-type lysozymes in a bivalve lysozyme phylogenetic tree predicted using Neighbor-Jointing method. Recombinantly expressed MmeLys showed lysozyme activity and strong antibacterial activity against Gram positive and Gram negative bacteria. MmeLys mRNA and protein were detected to be mainly produced in hepatopancreas and gill by the methods of semi-quantitative RT-PCR and western blotting. In addition, MmeLys gene expression increased following Vibrio parahaemolyticus challenge. Results of this research indicated that MmeLys represents a new i-type lysozyme that likely functions in M. meretrix immunity. 相似文献
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Cloning and characterization of the tiger shrimp lysozyme 总被引:2,自引:0,他引:2
Lysozymes are key proteins to invertebrates in the innate immune responses against bacterial infections. A lysozyme gene isolated
from tiger shrimp, Penaeus monodon, was cloned, sequenced and characterized. The cDNA consists of a signal peptide of 18 amino acids and a mature peptide of
140 amino acids. The lysozyme is presumed to be a chicken-type lysozyme for it possesses two catalytic sites and eight cysteine
residues which are highly conserved across species of chicken-type lysozymes. The lysozyme cDNAs of Penaeus semisulcatus, Litopenaeus vannamei, Macrobrachium nipponense and Macrobrachium rosenbergii were also cloned. High similarities existed among shrimp and prawn lysozymes but phylogenetic relationship of shrimps and
prawns based on lysozyme molecules did not quite consistent with traditional taxonomic classification. High mRNA expression
was detected in hepatopancreas, haemocytes and gill of tiger shrimp. Recombinant lysozyme exhibited potent lytic activities
against fish pathogens providing evidence of the involvement of lysozyme in shrimp immunity. 相似文献
3.
Phylogenetic Analysis of Invertebrate Lysozymes and the Evolution of Lysozyme Function 总被引:22,自引:0,他引:22
Bachali S Jager M Hassanin A Schoentgen F Jollès P Fiala-Medioni A Deutsch JS 《Journal of molecular evolution》2002,54(5):652-664
We isolated and sequenced the cDNAs coding for lysozymes of six bivalve species. Alignment and phylogenetic analysis showed
that, together with recently described bivalve lysozymes, the leech destabilase, and a number of putative proteins from extensive
genomic and cDNA analyses, they belong to the invertebrate type of lysozymes (i type), first described by Jollès and Jollès
(1975). We determined the genomic structure of the gene encoding the lysozyme of Mytilus edulis, the common mussel. We provide evidence that the central exon of this gene is homologous to the second exon of the chicken
lysozyme gene, belonging to the c type. We propose that the origin of this domain can be traced back in evolution to the origin
of bilaterian animals. Phylogenetic analysis suggests that i-type proteins form a monophyletic family.
Received: 21 May 2001 / Accepted: 22 October 2001 相似文献
4.
M.M. Costa M. Prado-Alvarez C. Gestal H. Li P. Roch B. Novoa A. Figueras 《Fish & shellfish immunology》2009,26(3):515-523
The effect of live bacteria (Micrococcus lysodeikticus and Vibrio anguillarum), and PAMPs (poly I:C, zymosan, LPS, LTA and CpG) on the production of intermediate toxic radicals (respiratory burst activity and production of nitric oxide) and mytilin B, myticin C and lysozyme gene expression was studied in vivo and in vitro. In vitro, bacteria were able to modulate the haemocytes' respiratory burst activity, being significantly increased after 6 h of incubation. The effect of pathogen-associated molecular patterns (PAMPs) was also studied. Zymosan produced an increase of the PMA-mediated response but an inhibition of the zymosan-mediated response. A significant increase of nitric oxide production was found at all the sampled time points (1, 3 and 6 h) in comparison with controls on both, the Gram-positive and Gram-negative bacteria. The in vivo responses measured on haemocytes after M. lysodeikticus injection were faster than those induced by V. anguillarum. However, V. anguillarum induced stronger in vitro effects. Mytilin B, myticin C and lysozyme in vitro gene expression, occurred at short times after infection. The maximum in vitro expression was detected 3 h post-infection. The differences between M. lysodeikticus and V. anguillarum in different measured parameters may suggest that different signalling pathways might be involved. Moreover, among all assayed PAMPs, LPS elicited the highest response. 相似文献
5.
Matsumoto T Nakamura AM Takahashi KG 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2006,145(3-4):325-330
In bivalve molluscs including oysters, lysozymes play an important role in the host defense mechanisms against invading microbes. However, it remains unclear in which sites/cells the lysozyme genes are expressed and which subsequently produced the enzyme. This study cloned lysozyme cDNAs from the digestive organs of Pacific oyster Crassostrea gigas and European flat oyster Ostrea edulis. Both complete sequences of two oysters' lysozymes were composed of 137 amino acids. Two translated proteins present a high content in cysteine residues. Phylogenetic analyses showed that these oysters' lysozymes clustered with the invertebrate-type lysozymes of other bivalve species. In the Pacific oyster, lysozyme mRNA was expressed in all tissues except for those of the adductor muscle. In situ hybridization analyses revealed that lysozyme mRNA was expressed strongly in basophil cells in the digestive gland tubule of C. gigas, but not in digestive cells. Results indicated that the basophil cells of the oyster digestive gland are the sites of lysozyme synthesis. 相似文献
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两种沼虾溶菌酶基因ORF的克隆和罗氏沼虾溶菌酶基因的组织表达(英文) 总被引:8,自引:0,他引:8
分别提取罗氏沼虾和日本沼虾血细胞总RNA,RT-PCR扩增获得特异性cDNA片段,纯化后克隆到T载体上。序列测定表明所克隆的两种沼虾溶菌酶基因的开放阅读框(ORF)为477bp,共编码158个氨基酸,包括溶菌酶成熟肽140个氨基酸残基和信号肽18个氨基酸残基。同源性分析表明,罗氏沼虾和日本沼虾溶菌酶基因的碱基序列及推测氨基酸序列高度同源,分别为99.4%和98.1%。两种沼虾溶菌酶基因的碱基序列和推测氨基酸序列与Gen-Bank上其他对虾溶菌酶的同源性达83.0%和80.0%以上。两种沼虾溶菌酶都具有c-型溶菌酶典型的两个酶活性位点(Glu51)和(Asp68),以及8个保守结构氨基酸残基Cys,且在101、106和107位上缺少Asp,因而推测本实验所克隆的两种沼虾溶菌酶基因属c-型溶菌酶基因的非钙结合亚型。以PCR法制备罗氏沼虾溶菌酶基因的生物素标记探针,斑点杂交检测感染弧菌后溶菌酶基因mRNA在各组织中的转录水平,结果表明受感染6h后在眼、肌肉、鳃、肝胰腺、肠管中的表达量均有升高,其中在肝胰腺中的表达量最高,约为对照组的560%。在不同感染时间里,肝胰腺中该基因表达量有较大的变化:感染后3h表达量最低,24h后表达量升至最高,大约为对照组的430%,48h时的表达量又有所下降,但仍明显高于对照组(约为330%)。受弧菌感染后罗氏沼虾溶菌酶基因转录的上调证明溶菌酶基因在非特异性免疫中的直接作用,同时表明肝胰腺可能在沼虾的免疫防御过程起重要作用。
相似文献
7.
Lysozymes in the animal kingdom 总被引:11,自引:0,他引:11
Lysozymes (EC 3.2.1.17) are hydrolytic enzymes, characterized by their ability to cleave the β-(1,4)-glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in peptidoglycan, the major bacterial cell wall polymer. In the animal kingdom, three major distinct lysozyme types have been identified — the c-type (chicken or conventional type), the g-type (goose-type) and the i-type (invertebrate type) lysozyme. Examination of the phylogenetic distribution of these lysozymes reveals that c-type lysozymes are predominantly present in the phylum of the Chordata and in different classes of the Arthropoda. Moreover, g-type lysozymes (or at least their corresponding genes) are found in members of the Chordata, as well as in some bivalve mollusks belonging to the invertebrates. In general, the latter animals are known to produce i-type lysozymes. Although the homology in primary structure for representatives of these three lysozyme types is limited, their three-dimensional structures show striking similarities. Nevertheless, some variation exists in their catalytic mechanisms and the genomic organization of their genes. Regarding their biological role, the widely recognized function of lysozymes is their contribution to antibacterial defence but, additionally, some lysozymes (belonging to different types) are known to function as digestive enzymes. 相似文献
8.
Purification and characterization of lysozyme from plasma of the eastern oyster (Crassostrea virginica) 总被引:1,自引:0,他引:1
Xue QG Schey KL Volety AK Chu FL La Peyre JF 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》2004,139(1):11-25
Lysozyme was purified from the plasma of eastern oysters (Crassostrea virginica) using a combination of ion exchange and gel filtration chromatographies. The molecular mass of purified lysozyme was estimated at 18.4 kDa by SDS-PAGE, and its isoelectric point was greater than 10. Mass spectrometric analysis of the purified enzyme revealed a high-sequence homology with i-type lysozymes. No similarity was found however between the N-terminal sequence of oyster plasma lysozyme and N-terminal sequences of other i-type lysozymes, suggesting that the N-terminal sequences of the i-type lysozymes may vary to a greater extent between species than reported in earlier studies. The optimal ionic strength, pH, cation concentrations, sea salt concentrations, and temperature for activity of the purified lysozyme were determined, as well as its temperature and pH stability. Purified oyster plasma lysozyme inhibited the growth of Gram-positive bacteria (e.g., Lactococcus garvieae, Enterococcus sp.) and Gram-negative bacteria (e.g., Escherichia coli, Vibrio vulnificus). This is a first report of a lysozyme purified from an oyster species and from the plasma of a bivalve mollusc. 相似文献
9.
Qinggang Xue Michael E Hellberg Kevin L Schey Naoki Itoh Ron I Eytan Richard K Cooper Jerome F La Peyre 《BMC evolutionary biology》2010,10(1):213
Background
Lysozymes are enzymes that lyse bacterial cell walls, an activity widely used for host defense but also modified in some instances for digestion. The biochemical and evolutionary changes between these different functional forms has been well-studied in the c-type lysozymes of vertebrates, but less so in the i-type lysozymes prevalent in most invertebrate animals. Some bivalve molluscs possess both defensive and digestive lysozymes. 相似文献10.
Invertebrate lysozymes: Diversity and distribution, molecular mechanism and in vivo function 总被引:1,自引:0,他引:1
Lysozymes are antibacterial enzymes widely distributed among organisms. Within the animal kingdom, mainly three major lysozyme types occur. Chicken (c)-type lysozyme and goose (g)-type lysozyme are predominantly, but not exclusively, found in vertebrate animals, while the invertebrate (i)-type lysozyme is typical for invertebrate organisms, and hence its name. Since their discovery in 1975, numerous research articles report on the identification of i-type lysozymes in a variety of invertebrate phyla. This review describes the current knowledge on i-type lysozymes, outlining their distribution, molecular mechanism and in vivo function taking the representative from Venerupis philippinarum (formerly Tapes japonica) (Vp-ilys) as a model. In addition, invertebrate g-type and ch-type (chalaropsis) lysozymes, which have been described in molluscs and nematodes, respectively, are also briefly discussed. 相似文献
11.
Chenghua Li Huili Sun Aiqin Chen Xuanxuan Ning Huifeng Wu Song Qin Qinzhao Xue Jianmin Zhao 《Fish & shellfish immunology》2010,28(3):499-503
Superoxide dismutase (SOD, EC 1.15.1.1) represents one kind of enzyme involved in scavenging the high level of reactive oxygen species (ROS) into molecular oxygen and hydrogen peroxide. In the present study, the intracellular Cu/Zn-SOD gene (icCu/Zn-SOD) of Venerupis philippinarum (denoted as VpSOD) was identified from haemocytes by homology cloning and RACR approaches. The full-length cDNA of VpSOD consisted of 910 nucleotides with a canonical polyadenylation signal sequence AATAAA, a polyA tail, and an open-reading frame of 465 bp encoding 154 amino acids. The deduced amino acid of VpSOD shared high similarity with the icCu/Zn-SODs from other species, indicating that VpSOD should be a new member of icCu/Zn-SOD family. Several highly conserved motifs including Cu, Zn binding sites (H46, H48, H63, H120 for Cu binding, and H63, H71, H80, D83 for Zn binding), intracellular disulfide bond and two Cu, Zn SOD signatures were also identified in VpSOD. The temporal expression of VpSOD in haemocytes after Vibrio anguillarum challenge was recorded by quantitative real-time RT-PCR. The relative expression level of VpSOD mRNA was up-regulated rapidly at 6 h post-infection and reached 18-fold of the control group. After a drastic decrease at 12 h, the expression level increased again and reached 22-fold to that in the control group at 96 h post-infection. All these results indicated that VpSOD was an acute-phase protein involved in the immune responses of V. philippinarum. 相似文献
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Boehnisch C Wong D Habig M Isermann K Michiels NK Roeder T May RC Schulenburg H 《PloS one》2011,6(9):e24619
Pathogens represent a universal threat to other living organisms. Most organisms express antimicrobial proteins and peptides, such as lysozymes, as a protection against these challenges. The nematode Caenorhabditis elegans harbours 15 phylogenetically diverse lysozyme genes, belonging to two distinct types, the protist- or Entamoeba-type (lys genes) and the invertebrate-type (ilys genes) lysozymes. In the present study we characterized the role of several protist-type lysozyme genes in defence against a nematocidal strain of the Gram-positive bacterium Bacillus thuringiensis. Based on microarray and subsequent qRT-PCR gene expression analysis, we identified protist-type lysozyme genes as one of the differentially transcribed gene classes after infection. A functional genetic analysis was performed for three of these genes, each belonging to a distinct evolutionary lineage within the protist-type lysozymes (lys-2, lys-5, and lys-7). Their knock-out led to decreased pathogen resistance in all three cases, while an increase in resistance was observed when two out of three tested genes were overexpressed in transgenic lines (lys-5, lys-7, but not lys-2). We conclude that the lysozyme genes lys-5, lys-7, and possibly lys-2 contribute to resistance against B. thuringiensis, thus highlighting the particular role of lysozymes in the nematode's defence against pathogens. 相似文献
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《Fish & shellfish immunology》2014,36(1):305-314
Lysozyme is an important component of the innate immune response against pathogen infection. The gene coding for c-type lysozyme in red-spotted grouper Epinephelus akaara was cloned and designated EaClys. The complete cDNA contains a 432 bp open reading frame encoding a protein of 144 amino acids displaying 65–91% similarity with the amino acid sequences of human, mouse, chicken, and fish counterparts. Recombinant EaClys (rEaClys) was expressed in Escherichia coli, displayed antibacterial activity against Gram-positive and Gram-negative bacteria, and possessed bactericidal activity against Vibrio alginolyticus. EaClys mRNA was constitutively expressed in all tested E. akaara tissues, and its expression increased after pathogen challenge. Most notably, challenges with LPS, SGIV or V. alginolyticus upregulated EaClys mRNA expression in the head, kidney, and blood. Its expression peaked between 16 and 24 h after challenge before dropping back to the baseline level. By using recombinant cytokines as signaling pathway mimetics and blocking antibodies and chemical inhibitors as pathway inhibitors, we show that LPS-induced lysozyme release from macrophages is promoted by cytokines TNF-α and IL-1β, and dependent on NF-κB pathway activation. These data suggest that EaClys is a constitutive and inducible acute-phase protein that is involved in the innate immune defense of E. akaara, and provide new clues about the molecular mechanisms that regulate innate immune responses in fish. 相似文献
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The genomic DNA of Escherichia coli, which contains the unmethylated CpG motif, was used to evaluate the immunostimulating effect of bacterial DNA on innate immune responses in the bivalve mussel Hyriopsis cumingii Lea. The results showed that the E. coli DNA had no significant effect on the production of superoxide anion (O(2-)) or acid phosphatase (AP) by haemocytes in vitro. However, the bactericidal activity of the haemocytes was significantly increased when the cells were incubated with 50 or 100mug/ml bacterial DNA for 12 and 24h. Antibacterial activity, lysozyme activity, and prophenoloxidase (proPO) production of haemolymph were also increased, when the bivalve molluscs were injected with 50 or 100mug/ml of bacterial DNA for 12 and 24h. These activities returned to the control level after 48h. This work showed the bacterial DNA with unmethylated CpG motif could activate some parameters of the immune system of bivalve molluscs in vivo and in vitro. 相似文献