Cloning, sequencing, and expression of a fibronectin/fibrinogen-binding protein from group A streptococci.

Lipoteichoic acid and several streptococcal proteins have been reported to bind fibronectin (Fn) or fibrinogen (Fgn), which may serve as host receptors. We searched for such proteins by screening a library of genes from M type 5 group A streptococci cloned into Escherichia coli. Lysates of clones were probed with biotinylated Fn and biotinylated Fgn. One clone expressed a 54-kDa protein that reacted with Fn and Fgn. The protein, termed FBP54, was purified and used to immunize rabbits. Anti-FBP54 serum reacted with purified, recombinant FBP54 and with a protein of similar electrophoretic mobility in extracts of M type 5, 6, and 24 streptococci. Anti-FBP54 serum also reacted with 5 of 15 strains of intact, live streptococci, suggesting that FBP54 may be a surface antigen. Southern blot analysis confirmed that the gene is found in group A streptococci but not in Staphylococcus aureus or E. coli. The cloned gene was sequenced and contained an open reading frame encoding a protein with a calculated molecular weight of 54,186. Partial amino acid sequencing of purified FBP54 confirmed that this open reading frame encoded the protein. As determined by utilizing fusion proteins containing truncated forms of FBP54, the primary Fn/Fgn-binding domain appears to be contained in residues 1 to 89. These data suggest that FBP54 may be a surface protein of streptococci that reacts with both Fn and Fgn and therefore may participate in the adhesion of group A streptococci to host cells.     

短穗鱼尾葵花粉泛过敏原profilin基因的克隆表达、纯化及免疫学鉴定

目的克隆并表达短穗鱼尾葵花粉中泛变应原肌动蛋白抑制蛋白（profilin）。方法利用RT-PCR结合RACE技术克隆短穗鱼尾葵花粉中泛变应原profilin的全长基因,并进行序列分析。然后设计带有酶切位点的特异性引物,采用RT-PCR获得整个短穗鱼尾葵花粉profilin的开放阅读框,将其与pET28a载体连接并转化大肠杆菌BL21（DE3）进行诱导表达,通过Ni2＋亲和层析柱对重组蛋白进行纯化,采用Western-blot检测其IgE结合活性。结果克隆获得了短穗鱼尾葵花粉profilin的全长基因,由608个碱基组成,开放阅读框为396个碱基（包括终止密码子）,编码131个氨基酸。该序列编码的蛋白为小分子量酸性蛋白,等电点为4.52,相对分子质量约为14200。此序列已被GenBank收录,登陆号为EF173600。重组短穗鱼尾葵花粉profilin在大肠杆菌中高效的表达,进一步经Ni2＋亲和层析柱纯化后经Western-blot检测具有良好的免疫学活性。结论成功地克隆和表达了短穗鱼尾葵花粉profilin,为短穗鱼尾葵花粉过敏的诊断和免疫治疗奠定了基础。     

Cloning and expression of the dnaK gene of Campylobacter jejuni and antigenicity of heat shock protein 70

Campylobacter jejuni is a leading cause of infectious diarrhea throughout the world. In addition, there is growing evidence that Guillain-Barré syndrome, an inflammatory demyelinating disease of the peripheral nervous system, is frequently preceded by C. jejuni infection. In the present study, the hrcA-grpE-dnaK gene cluster of C. jejuni was cloned and sequenced. The dnaK gene consists of an open reading frame of 1,869 bp and encodes a protein with a high degree of homology to other bacterial 70-kDa heat shock proteins (HSPs). The overall percentages of identity to the HSP70 proteins of Helicobacter pylori, Borrelia burgdorferi, Chlamydia trachomatis, and Bacillus subtilis were calculated to be 78.1, 60.5, 57.2, and 53. 8%, respectively. Regions similar to the Escherichia coli sigma70 promoter consensus sequence and to a cis-acting regulatory element (CIRCE) are located upstream of the hrcA gene. Following heat shock, a rapid increase of dnaK mRNA was detectable, which reached its maximum after 20 to 30 min. A 6-His-tagged recombinant DnaK protein (rCjDnaK-His) was generated in E. coli, after cloning of the dnaK coding region into pET-22b(+), and purified by affinity and gel filtration chromatography. Antibody responses to rCjDnaK-His were significantly elevated, compared to those of healthy individuals, in about one-third of the serum specimens obtained from C. jejuni enteritis patients.     

Cloning of the glutamine synthetase gene from group B streptococci.

The glnA gene from the human pathogen Streptococcus agalactiae was cloned from a genomic library prepared with the lambda phage vector lambdaDASHII. A 4.6-kb DNA fragment of one of the recombinant phages was subcloned in pUC18. This Escherichia coli clone expressed a 52-kDa protein encoded by a 1,341-bp open reading frame. The nucleotide sequence of the open reading frame and the deduced amino acid sequence shared a significant degree of homology with the sequences of other glutamine synthetases (GS). The highest homology was between our deduced protein and GS of gram-positive bacteria such as Bacillus subtilis, Bacillus cereus, and Staphylococcus aureus. Plasmids with the cloned streptococcal glnA were able to complement E. coli glnA mutants grown on minimal media. Rabbit antisera to streptococcal GS recombinant protein recognized not only the recombinant protein but also a similar-sized band in mutanolysin extracts of all group B streptococcal strains tested, regardless of polysaccharide type or surface protein profile. The amino acid sequence of the deduced protein had similarities to other streptococcal cell-surface-bound proteins. The possible functional role of the immunological features of streptococcal GS is discussed.     

The 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide is a DnaK-like protein.

The gene coding for the 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide has been cloned in Escherichia coli, and the nucleotide sequence has been determined. The cloned DNA fragment contained the coding region as well as the putative promoter. The deduced amino acid sequence of the 1,980-base-pair open reading frame revealed 94% homology with a 75-kilodalton protein from C. trachomatis serovar D and 57% homology with the DnaK proteins of E. coli and of Bacillus megaterium, while amino acid homology with human heat shock protein 70 (hsp70) was 42%. The promoter region was identified by computer search and by primer extension of mRNA synthesized in recombinant E. coli. The promoter region which differed from the putative promoter region in serovar D was shown to be a mixed promoter type in which the -10 region showed a regular TATA box configuration while the -35 region showed high homology with heat shock promoters. This mixed promoter was recognized in E. coli.     

Characterization of a novel Candida albicans 29 kDa IgE-binding protein--purification, cDNA isolation and heterologous expression of Cand a 3

BACKGROUND: Candida albicans has been implicated in human allergic disorders. However, many of its immunoglobulin E (IgE)-reacting components have not yet been identified. The purpose of the present study is to characterize a novel 29 kDa IgE-binding protein from C. albicans. METHODS: The 29 kDa protein was partially purified and its tryptic digests subjected to mass spectrometric analysis. The cDNA encoding this protein was isolated and heterologously expressed in Escherichia coli. Monoclonal antibodies (MoAbs) were raised against the 29 kDa protein purified from C. abicans extracts. RESULTS: We isolated a 29 kDa IgE-reacting component from C. albicans. The protein was digested on-gel with trypsin and the masses of the resulting fragments were determined in a MALDI-TOF mass spectrometer. The data were searched against protein sequences deduced from the C. albicans genome. An open reading frame that possibly encodes the 29 kDa IgE-reacting component was identified. The cDNA corresponding to the open reading frame was isolated. It encodes a 236 residues protein that has 62% sequence identity to that of a hypothetical protein (YDR533c) from Saccharomyces cerevisiae. Conserved domain search suggests that the encoded protein belongs to the ThiJ/PfpI family. The cDNA isolated was inserted into a pQE-30 vector for protein expression in Escherichia coli. The recombinant protein can react with IgE antibodies in sera from asthmatic patients and two MoAbs that were generated against the purified native 29 kDa protein from C. albicans. CONCLUSIONS: We identified and cloned a novel 29 kDa IgE-reacting component (Cand a 3) from C. albicans. The recombinant proteins produced from this clone and the MoAbs prepared may be useful in the standardization of diagnostic extracts. They are also instrumental in elucidating the role of C. albicans in clinical allergy.     

A simple method for the production of specific antiserum to protein encoded in cloned genes. Immunization with precipitin lines

A simple technique for raising specific antisera to protein encoded by cloned genes is described. The procedure involves preparation of an antiserum to Escherichia coli beta-galactosidase and the use of that serum to immunoprecipitate a fusion protein in a crossed immunoelectrophoresis gel followed by immunization with fusion protein precipitin arcs. An antiserum was prepared against protein encoded by an open reading frame in a dispersed repeated DNA sequence found in the protozoan Trypanosoma brucei. This serum recognized a polypeptide doublet of 33.5 and 32.5 kDa on immunoblots prepared from extracts of T. brucei. The method described should be applicable to other investigations where an immunochemical reagent against protein encoded by a cloned gene is desired.     

Detection of humoral response using a recombinant heat shock protein 70, DnaK, of Mycoplasma haemofelis in experimentally and naturally hemoplasma-infected cats

Hemoplasmas is the trivial name given to a group of erythrocyte-parasitizing bacteria of the genus Mycoplasma. Of the feline hemoplasmas, Mycoplasma haemofelis is the most pathogenic, while "Candidatus Mycoplasma haemominutum" and "Candidatus Mycoplasma turicensis" are less pathogenic. Shotgun libraries of fragmented M. haemofelis genomic DNA were constructed, and random colonies were selected for DNA sequencing. In silico-translated amino acid sequences of putative open reading frames were compared to mass spectrometry data from M. haemofelis protein spots identified as being immunogenic by two-dimensional gel electrophoresis and Western blotting. Three of the spots matched the predicted sequences of a heat shock protein 70 (DnaK) homolog, elongation factor Ts, and a fragment of phosphoglycerate kinase found during library screening. A full-length copy of the M. haemofelis dnaK gene was cloned into Escherichia coli and recombinantly expressed. Recombinant M. haemofelis DnaK was purified and then used in Western blotting and an enzyme-linked immunosorbent assay (ELISA) to investigate the humoral immune response during acute infection in cats experimentally infected with M. haemofelis, "Ca. Mycoplasma haemominutum," or "Ca. Mycoplasma turicensis". The recombinant M. haemofelis DnaK ELISA also was used to screen clinical samples submitted for hemoplasma PCR testing to a commercial laboratory (n = 254). Experimentally infected cats became seropositive following infection, with a greater and earlier antibody response seen in cats inoculated with M. haemofelis than those seen in cats inoculated with "Ca. Mycoplasma haemominutum" or "Ca. Mycoplasma turicensis," by both Western blotting and ELISA. Of the clinical samples, 31.1% had antibodies detected by the ELISA but only 9.8% were positive by PCR for one or more hemoplasmas.     

The proteinase polypeptide of adenovirus serotype 2 virions

The Ad2 proteinase, which is thought to be encoded by a 23-kDa open reading frame located at the end of the L3 family of late mRNAs, is expressed poorly even late after infection. To obtain sufficient proteinase for biochemical characterization, a DNA fragment containing the 23-kDa open reading frame was cloned into plasmids that permit efficient expression in Escherichia coli. Polyclonal antiserum specific for the Ad2 proteinase was produced by immunizing rabbits with a fusion protein that included the entire proteinase open reading frame, and this antiserum was used to show that the product of the 23-kDa reading frame is assembled into virions. Bacterial products corresponding to the complete 204 amino acid proteinase reading frame, to a 9 amino acid proteinase deletion, and to a proteinase fusion protein of 227 amino acids were used to determine the size of the proteinase polypeptide in Ad2 virions and in infected HeLa cell extracts. A single proteinase polypeptide that migrated during SDS-polyacrylamide gel electrophoresis with the 204 amino acid recombinant proteinase was detected in wild-type and H2ts1 virions, and in infected cell extracts. Immunoblot titrations showed that a wild-type Ad2 virus particle contains about 10 proteinase polypeptides; an H2ts1 virion has approximately fivefold less proteinase. In virions, the proteinase was associated primarily with the virus core. The 204 amino acid proteinase produced in E. coli permitted cleavage of the major core protein precursor, P-VII, to mature, authentic VII, but the proteinase deletion lacking 9 amino acids from near the amino-terminus was inactive. These results are inconsistent with autocatalytic processing of the Ad2 proteinase as was reported by Chatterjee and Flint (1987, Proc. Natl. Acad. Sci. USA 84, 714-718).     

一种华支睾吸虫亲肌素样基因的克隆、表达与免疫学功能初步研究

目的对新发现的华支睾吸虫亲肌素样蛋白（CsMLP）进行克隆、原核表达,初步了解其重组产物的免疫学功能。方法应用生物信息学分析软件,分析CsMLP的序列特点和基本理化特征;将CsMLP基因克隆到原核表达质粒pET．28a（＋）中,诱导表达并用镍离子金属螯合剂亲和层析柱进行纯化,用纯化的CsMLP蛋白免疫SD大鼠获得抗血清;用Westernblot进行免疫反应性及免疫原性分析;应用免疫荧光方法观察CsMLP在华支睾吸虫成虫的定位。结果该cDNA序列全长为900bp,编码190个氨基酸,具有Calponin功能域;PCR、双酶切及DNA测序结果均表明pET．28a（＋）-CsMLP重组质粒构建成功;SDS—PAGE结果表明目的基因在大肠杆菌BL21／DE3中获得高效表达,分子量为21300Mr;经亲和层析获得了高纯度蛋白;重组蛋白可被其免疫的SD大鼠血清、感染了华支睾吸虫的SD大鼠血清识别;CsMLP主要定位于虫体富含肌肉组织的口、腹吸盘、咽部,在表膜、肠壁也有分布。结论CsMLP可在原核表达系统中呈现高效的可溶性表达,具有免疫原性．其主要分布在华支睾吸虫肌肉丰富的组织。     

Nucleotide sequence of htpB, the Legionella pneumophila gene encoding the 58-kilodalton (kDa) common antigen, formerly designated the 60-kDa common antigen.

Gene htpB, which encodes the 58-kilodalton protein of Legionella pneumophila, was cloned in Escherichia coli and its complete nucleotide sequence was determined. Analysis of this sequence revealed an open reading frame of 1,644 nucleotides encoding a protein with a predicted molecular mass of 57,952 daltons. Data obtained by amino-terminal sequencing of the purified 58-kilodalton protein agreed, except for one amino acid residue, with the predicted amino acid sequence, identifying this open reading frame as htpB. A comparison of the primary structure of this protein to other proteins of similar molecular weights from E. coli, Mycobacterium leprae, M. tuberculosis, and Coxiella burnetii revealed significant regions of sequence similarity, which are discussed.     

Genetic analysis of the gene cluster encoding nonfimbrial adhesin I from an Escherichia coli uropathogen.

The chromosomally encoded nonfimbrial adhesion I (NFA-I) from Escherichia coli urinary tract isolate 827 (O83:K1:H4) mediates agglutination of human erythrocytes. Subclones were constructed from an NFA-I-expressing recombinant E. coli K-12 clone, derived from a genomic library of E. coli 827. Minicell analysis and nucleotide sequencing revealed that proteins of 30.5, 9, 80, 15, and 19 kDa encoded on a stretch of approximately 6 kb are involved in the expression of NFA-I. NFA-I exhibits a polymeric structure, which disintegrates with elevated temperature into a 19-kDa monomer but with some relatively stable dimers. By using gold-conjugated monoclonal antibodies directed against NFA-I in electron microscopy, the adhesin could be localized on the outer surface of the recombinant E. coli K-12 bacteria. The nucleotide sequence of the nfaA gene encoding the monomeric structural subunit of the adhesin was determined. An open reading frame of 184 amino acids encoding the NfaA precursor, which is processed to the mature protein, was found; it consisted of 156 amino acids with a calculated molecular weight of 16,000. Peptide sequencing of the NFA-I subunit protein confirmed that this open reading frame corresponds to the NfaA coding locus. Furthermore, the nucleotide sequence of the open reading frame termed NfaE, located at the proximal part of the DNA stretch responsible for NFA-I expression, was elaborated. NfaE consists of 247 amino acids, including a presumptive 29-amino-acid signal peptide, leading to a molecular weight of 24,000 for the mature protein. The nfaE sequence shares homology with the 27-kDa CS3 protein, which is involved in the assembly of CS3 fibrillae, and might encode the 30.5-kDa protein, detected in minicells.     

Molecular cloning and nucleotide sequence of the alpha-toxin (phospholipase C) of Clostridium perfringens.

A fragment of DNA containing the gene coding for the phospholipase C (alpha-toxin) of Clostridium perfringens was cloned into Escherichia coli. The cloned DNA appeared to code only for the alpha-toxin and contained both the coding region and its associated gene promoter. The nucleotide sequence of the cloned DNA was determined, and an open reading frame was identified which encoded a protein with a molecular weight of 42,528. By comparison of the gene sequence with the N-terminal amino acid sequence of the protein, a 28-amino-acid signal sequence was identified. The gene promoter showed considerable homology with the E. coli sigma 55 consensus promoter sequences, and this may explain why the gene was expressed by E. coli. The cloned gene product appeared to be virtually identical to the native protein. A 77-amino-acid stretch that was close to the N terminus of the alpha-toxin showed considerable homology with similarly located regions of the Bacillus cereus phosphatidylcholine, preferring phospholipase C and weaker homology with the phospholipase C from Pseudomonas aeruginosa.     

The Borrelia burgdorferi flagellum-associated 41-kilodalton antigen (flagellin): molecular cloning, expression, and amplification of the gene.

Monoclonal antibodies directed against the major Borrelia burgdorferi flagellar protein, the 41-kilodalton (kDa) protein flagellin, were used to monitor cloning and expression of the flagellin gene from a Borrelia burgdorferi genomic library. The structure of the gene was analyzed, and recombinant nonfusion flagellin was produced in Escherichia coli. A DNA sequence analysis of the 41-kDa flagellin gene revealed the presence of an open reading frame that encoded a protein having 336 amino acid residues and a calculated molecular mass of 35.8 kDa, indicating that there was posttranslational modification of the natural 41-kDa flagellin protein. Upstream from the AUG start codon sequence we identified motifs corresponding to consensus procaryotic promoter elements which could be utilized by the cloned flagellin gene when it was expressed in E. coli MC1061. The deduced flagellin protein sequence exhibited high levels of homology to sequences of flagellin proteins from Bacillus subtilis and Salmonella typhimurium. The levels of sequence similarity for the amino- and carboxy-terminal portions were about 65 and 56%, respectively. DNA sequence information on the flagellin gene was used to design oligonucleotides for gene amplification by the polymerase chain reaction method, and by using this method 0.01 pg of Borrelia burgdorferi DNA could be detected. Our results provide a basis for further biochemical analysis of the 41-kDa flagellin protein, investigation of the role of this protein in host-pathogen interactions, and development of a standardized reagent for diagnostic systems for Borrelia burgdorferi infections.     

金黄色葡萄球菌肠毒素A基因的克隆及原核表达

目的从分离培养的金黄色葡萄球菌中提取肠毒素A（SEA）基因,亚克隆入表达载体pET-28a（＋）,诱导融合蛋白的表达,为肠毒素A应用研究奠定基础。方法根据GenBank中SEA的序列,设计一对特异性引物,以金黄色葡萄球菌DNA为模板PCR扩增SEA,纯化DNA进行BamHⅠ、XholⅠ双酶切鉴定及测序鉴定,并与做相应酶切的pET-28a（＋）连接,转化大肠杆菌BL21,并对质粒进行双酶切鉴定及基因序列分析,用IPTG诱导融合蛋白的表达,His标签单克隆抗体进行免疫印迹验证融合蛋白的表达。结果以金黄色葡萄球菌DNA为模板,成功扩增了SEA基因,基因大小为774bp,重组PET-28a（＋）-SEA双酶切鉴定可见目的片段,测序结果显示SEA在正确读框中,序列比对分析显示其与相关报道核苷酸序列一致性达99.9%。经IPTG诱导后,SDS-PAGE可见pET-28a（＋）-SEA/BL21在相应分子量（约30000Mr）条带大量表达,免疫印迹能检测到目的蛋白条带,说明其与His标签融合表达。结论克隆了金黄色葡萄球菌SEA基因,并成功在大肠杆菌BL21中融合表达,为肠毒素A应用研究奠定了基础。     

Molecular characterization of the pathogen-specific, 34-kilodalton membrane immunogen of Treponema pallidum.

The 34-kilodalton (kDa) antigen of Treponema pallidum subsp. pallidum (T. pallidum) is a pathogen-specific integral membrane protein. DNA sequence analysis of the cloned gene revealed an open reading frame encoding a primary product of 204 residues with a molecular mass of 22,087 daltons. Sequences that correspond to a consensus Escherichia coli promoter and a ribosome-binding site were found upstream from the AUG start codon that begins the open reading frame, suggesting that the cloned gene can use its own regulatory sequences for expression. Examination of the deduced amino acid sequence revealed the presence of a typical procaryotic leader peptide 19 amino acids long; processing results in a mature molecule with a molecular mass of 20,123 daltons. Pulse-chase experiments with E. coli minicells confirmed that the 34-kDa antigen is synthesized as a higher-molecular-weight precursor that is processed to a mature form with the electrophoretic mobility that is characteristic for this protein. The presence in the leader peptide of the sequence Phe-Ser-Ala-Cys suggested that the 34-kDa antigen is a proteolipid. Although hydropathy analysis of the deduced amino acid sequence of the mature 34-kDa antigen predicted that the molecule was primarily hydrophilic, both the native and recombinant 34-kDa molecules displayed hydrophobic biochemical behavior by fractionating into the detergent phase after extraction of intact organisms with Triton X-114. Cell fractionation experiments with E. coli showed that the 34-kDa molecule was localized in both the inner and outer membranes of the recombinant host. The combined data demonstrate that the 34-kDa antigen is an integral membrane protein that behaves in a biochemically consistent manner in both T. pallidum and E. coli.     

鸡柔嫩艾美耳球虫江苏分离株5401基因的克隆、序列分析及原核表达

分别以孢子化卵囊及第2代裂殖子为材料,应用RT-PCR技术克隆了柔嫩艾美耳球虫（Eimeria tenella）江苏分离株5401基因。测序结果表明该序列全长为864bp,序列本身是一个开放阅读框,将克隆得到的基因与国外报道的5401基因比较,第174位和第287位发生突变,碱基分别由T突变为C,由C突变为T。第1个为无义突变,第2个突变引起第96位氨基酸由A变为V。其核苷酸序列同源性为99.8%,氨基酸同源性为99.7%。将所获5401基因克隆到pGEX-4T-2获得重组质粒pGEX-4T-2-5401,转入宿主菌BL21中,用IPTG进行诱导表达。用大鼠抗柔嫩艾美耳球虫子孢子高免血清对原核表达产物进行Western Blotting检测,结果表明有2条融合蛋白,大小分别为90kDa和80kDa左右。     

High-level expression of a 56-kilodalton protein gene (bor56) of Rickettsia tsutsugamushi Boryong and its application to enzyme-linked immunosorbent assays.

The 56-kDa protein of Rickettsia tsutsugamushi, which is located on the rickettsial surface, has been shown to be an immunodominant antigen. The gene that encodes the 56-kDa protein of R. tsutsugamushi Boryong (bor56) was cloned. Sequencing revealed an open reading frame of 1,602 bp encoding 534 amino acids with a molecular weight of 56,803. The 56-kDa protein of R. tsutsugamushi Boryong (Bor56) was expressed as a fusion protein with the maltose-binding protein of Escherichia coli by deleting 252 bp from the 5' end of the open reading frame and subcloning it into the StuI site of pIH821. The recombinant fusion protein was purified by amylose column chromatography for application in an enzyme-linked immunosorbent assay to evaluate the ability of the method to detect the antibody to R. tsutsugamushi in human patient sera. By using sera from 100 patients with scrub typhus and 70 patients with other febrile diseases, a high diagnostic sensitivity (95%) and a high diagnostic specificity (100%) were demonstrated, suggesting the suitability of the recombinant antigen for use as an immunodiagnostic tool.     

质粒蛋白pORF5在沙眼衣原体感染细胞中的定位及免疫原性研究(英文)

目的分析沙眼衣原体隐蔽性质粒编码的质粒蛋白pORF5在感染细胞中的定位并初步探讨免疫原性特征。方法 PCR扩增pORF5质粒蛋白编码基因,构建原核表达重组体pGEX-6p/pORF5,重组体转化大肠杆菌XL1-blue中诱导表达融合蛋白;融合蛋白经Glutathione Sepharose亲和层析纯化后免疫小鼠,制备单克隆抗体和多克隆抗体,间接免疫荧光技术及免疫印迹鉴定抗体的特异性,并分析pORF5质粒蛋白在感染细胞中的分布特征。采用ELISA方法分析pORF5质粒蛋白的免疫原性。结果 pORF5原核表达重组体成功构建,融合蛋白在大肠杆菌中可溶性表达;pORF5主要分布于宿主细胞胞浆,但也少量分布在EB、RB上;pORF5能与衣原体患者血清及鼠免疫血清发生强烈地免疫反应。结论 pORF5为衣原体分泌蛋白,并具有很强的免疫原性。