食用菌中HIV-1反转录酶和蛋白酶抑制剂的分离与纯化
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摘要
获得性免疫缺陷综合症(艾滋病)是一种因感染到人类免疫缺陷病毒(HIV)而导致的传染性疾病。人免疫缺陷病毒的反转录酶、蛋白酶和整合酶在病毒复制过程中起着十分重要的作用，缺一不可。目前较为有效的抗病毒化学治疗方法是反转录酶和蛋白酶抑制剂的复合用药，这些药物虽然能够对HIV有一定的抑制效果，但长期服用会对患者产生很大的毒副作用，且容易使病毒产生耐药性，所以需要寻找出毒性低且价格便宜的药物来对抗病毒。本论文测定了多种食用菌多糖复合物对反转录酶(RT)和蛋白酶(PR)的抑制作用。实验结果表明，对HIV-1 RT有抑制作用的组分主要集中在乙醇沉淀部分，即多糖复合物部分。其中以茯苓的活性最高，1mg／ml的反应浓度下抑制率达97.4％，其次是香栓菌90.6％和茶树菇84.5％。选取活性最高的茯苓菌丝体多糖复合物来作为纯化对象，经过DEAE-纤维素阴离子交换层析，CM-纤维素阳离子交换层析和FPLC Superdex 75凝胶过滤方法分离纯化后，获得一种对HIV-1 RT具有高抑制活性的糖蛋白S12，并测出其蛋白部分N末端的5个氨基酸顺序：丙氨酸—谷氨酸—天冬酰胺—甘氨酸—甘氨酸。在已测的22种食用菌多糖复合物对HIV-1 PR的抑制活性实验中，反应浓度为0.5mg／ml下对蛋白酶抑制活性最高的是香乳菇，其抑制率达80.9％。
Acquired Immunodeficiency Syndrome (AIDS) is a deadly pandemic disease caused by a retrovirus known as human immunodeficiency virus type 1(HIV-1). Reverse transcriptase (RT), protease (PR), and integrase (IN) are three essential enzymes to the life cycle of the HIV. The chemotherapeutic strategies have been directed at the development of inhibitors of these HIV enzymes. One of the major obstacles to the long-term treatment of AIDS is the remarkable mutation of HIV to most, if not all, of clinically used chemotherapeutic agents. Therefore, it is an ongoing need for new, structurally diverse anti-HIV agents, which are less toxic and affordable to all AIDS patients. In this study, we firstly screened the ethanol extracts from edible mushrooms on inhibition of HIV-1 RT activity. Among the mushroom ethanol extracts examined, the extract of Poria cocos exhibited over 97.4% inhibition at Img/ml. Then this crude ethanol extract was fractionated through DEAE-cellulose anion-exchange chromatography, CM-cellulose catio
n-exchange chromatography, and FPLC Superdex 75 gel filtration respectively, which result in the isolation of an active polysaccharide-peptide with five amino acids of AENGG in N-terminal, named SI 2. On the other hand, we also examined these extracts' inhibitory effects on HIV-1 PR. The results showed that Lactarius camphorates, which could inhibit HIV-1 PR by 80.9%, was the only sample exhibited over 80% inhibition on PR at 0.5mg/ml.
Acquired Immunodeficiency Syndrome (AIDS) is a deadly pandemic disease caused by a retrovirus known as human immunodeficiency virus type 1(HIV-1). Reverse transcriptase (RT), protease (PR), and integrase (IN) are three essential enzymes to the life cycle of the HIV. The chemotherapeutic strategies have been directed at the development of inhibitors of these HIV enzymes. One of the major obstacles to the long-term treatment of AIDS is the remarkable mutation of HIV to most, if not all, of clinically used chemotherapeutic agents. Therefore, it is an ongoing need for new, structurally diverse anti-HIV agents, which are less toxic and affordable to all AIDS patients. In this study, we firstly screened the ethanol extracts from edible mushrooms on inhibition of HIV-1 RT activity. Among the mushroom ethanol extracts examined, the extract of Poria cocos exhibited over 97.4% inhibition at Img/ml. Then this crude ethanol extract was fractionated through DEAE-cellulose anion-exchange chromatography, CM-cellulose catio
n-exchange chromatography, and FPLC Superdex 75 gel filtration respectively, which result in the isolation of an active polysaccharide-peptide with five amino acids of AENGG in N-terminal, named SI 2. On the other hand, we also examined these extracts' inhibitory effects on HIV-1 PR. The results showed that Lactarius camphorates, which could inhibit HIV-1 PR by 80.9%, was the only sample exhibited over 80% inhibition on PR at 0.5mg/ml.
引文
Andrake M.D and Skalka A.M. Retrovial integrase, putting the pieces together. Journal of Biological Chemistry, 1996, 271:19633～36
Anthony S. Fauci. The human immunodeficiency virus: infectivity and mechanisms of pathogenesis. Science, 1988, 239:617～622
Bagarazzi M.I., Boyer J.D. and Ayyavoo V. Nucleic acid-based vaccines as an approach to immunization against human immunodeficiency virus type-Ⅰ. Current Top Microbiological Immunology,1998, 226:107～143
Barreiro P., Soriana V. and Blanco F. Risks and benefits of replacing protease inhibitors by nevirapine in HIV-infected subjects under long-term successful triple combination therapy. AIDS, 2000, 14:807～812
Bartlett J.G. Protease inhibitor for HIV infection. Annals of Internal Medicine, 1996, 124: 1086～1087
Cai M., Huang Y., Caffrey M., Zheng R., Craigie R. and Clore GM. Solution structure of the N-terminal zinc binding domain of HIV integrase. Nature Structural Biology. 1997, 4:567～577
Chang R.S. and Yeung H.W. Inhibiton of growth of human immunodeficiency virus in vitro by crude extracts of Chinese medicinal herbs. Antiviral Research, 1988, 9:163～176
Chang S.T. and Miles P.G. Mushroom biology-A new discipline. The Mycologist, 1992, 6:64～65
Chihara G., Hamuro J., Maeda Y.Y., Ami Y. and Fukuoka F. Fraction and Purification of the polysaccharides with marked antitumor activity, especially Lentinan from Lentinus edodes. Cancer Research, 1970, 30:943～944
Chung R.S. Functional properties of edible mushroom. Nutrition Review, 1996, 54(11):91～94
Chopra K.F. and Tyring S.K. Current antiretroviral therapy in the treatment of HIV infection. Seminars Cutaneou Medical Surgery, 1997, 16:381～382
Collins R.A., Ng T.B., Fong W.P., Wan C.C. and Yeung H.W. A comparison of human immunodeficiency virus type Ⅰ infection by partially purified aqueous extracts of Chinese medicinal herbs. Life Sciences, 1997, 60:pc345～351
Cooper J.I. The nature of virus. In Virus and Environment(2nd ed.). London: Chapman & Hall, 1995, 1～33
Dalgeish A.G. and Beverly P.L. The CD4(T4) antigen is an essential compnent of the receptor of the AIDS retrovirus. Nature, 1984, 312:763～767
Debyser Z. HIV-1 Reverse transcription in viral DNA synthesis. Leuven: Leuven University Press. 1994, 23～24
Emernan M. and Malim M.H. HIV-1 regulatory/accessory genes: key to unraveling viral and host cell biology. Science, 1998, 280:1880～1884
Fesen M.R., Kohn K.W., Leteurtre F. and Pommier Y. Inhibitors of human immunodeficiency virus integrase. Proceedings of National Academy of Sciences of the USA, 1993, 90:2399～2340
Fisher R.J., Rein A., Fivashi M., Urbaneja M.A., Casas-finet J.R., Medaglia M. and Henderson L.E. Sequence-specific binding of human immunodeficiency virus type Ⅰ nucleacapsid protein to shoot oligonucleotides. Journal of Virology. 1998, 72:1902～1909
Francesco M.A., Caruso A., Fallacara F., Canaris A., Francesco D., Poiesi C., Licenziati S., Coalli M., Martinielli F., Fiorentini S. and Turano A. HIV p17 enhances lymphocyte proliferation and HIV-I replication after binding to a human serum factor. AIDS. 1998, 12:245～252
Gerton J.L., Sharron O., Mari O., Joseph D. and Patrick D. Effects of mutations in residues near the active site of human immunodeficiency virus type Ⅰ integrase on specific enzyme-substrate interaction. Journal of Virology. 1998, 72:5046～5055
Gotch F., Hardy G. and Imami N. Therapeutic vaccines in HIV-1 infection. Immunology Review, 1999,170:173～182
Goto T. and Komuro N. Projection structures of human immunodeficiency virus type 1(HIV-1) observed with high resolution electron microscopy. Journal of Electron Microscopy. 1994, 43:16～19
He J.L. CCR3 and CCR5 are co-receptors for HIV-1 infection of microgha. Nature. 1997, 385:645～649
Hammer S.M. Antiretroviral therapy: where are we? AIDS. 1998, 12:S181～S188
Hermann T., Meirer T., Gotte M. and Heumann H. The 'helix clamp' in HIV-1 reverse transeriptase nuleic acid binding motif common in nucleic acid polymerase. Nucleic Acid Research, 1994, 22:1686～1695
Hoffman T.L. Chemokines and co-receptors in HIV/SIV-host interaction. AIDS, 1998, S17～S26
Huang M.J. Incorporation of p160~(gag-pol) into virus particles requires the presence of both the major homology region and adjacent C-terminal capsid sequences with the Gag-Pol polyprotein. Journal of Virology. 1997, 4412～4418
Ikekawa T., Cehara N., Maeda Y., Nakanishi M. and Fukuoka E Antitumor activity of aqueous extracts of edible mushrooms. Cancer Research, 1969, 29:734～735
Jing N., Marchand C., Liu J., Mitra R., Hogan M.E. and Pommier Yves. Mechanism of inhibition of HIV-1 integrase by G-tetrad-forming oligonucleotides in vitro. Journal of Biological Chemistry. 2000, 275:21460～21467
Johnston M.I. HIV vaccines: problems and prospects. Hosp. Prac. (OffS. Ed.), 1997, 32:125～140
Kaneda T., Arai K. and Tokuda S. The effect of dried mushroom: Cortinellus shiitake on cholesterol metabolism in rats. Journal of Japanese Social Food Nutrition, 1964, 16:466～468
Khan E., Mack J.P., Katx R.A., Kullkosky J. and Skalka A.M. Retroviral integrase domains: DNA binding and the recognition of LTP sequences. Nucleic Acids Research, 1991, 19:581～860
Kraus E., James W. and Barclay A.N. Cutting Edge: noval RNA ligands able to bind CD4 antigen and inhibit CD4~+ T lymphocyte function. The journal of immunology. 1998, 160:5309～5212
Lafeuillade A. Effects of a comvination fo zidovudine, didanosine and lamivudine on primary human immunodeficiency virus type 1 infection. Journal of Infectious Disease. 1997,175～124
Levy J.A. HIV and the pathogenesis of AIDS. ASM Press, 1994
Littvak S. Retroviral Reverse Transcriptase. R. G. Lands Company, 1996
Luban J. Absconding with the chaperone: essential cyclophilin-gag interaction in HIV-1 virion. Cell. 1996, 87:1157～1159
Luciw P.A. Human immunodeficiency virus and their replication. In Field B.N. et al. ed Field Virology(3rd ed.). Philadelphia: Lippincott-Raven Publishers, 1996, 1881～1952
Mazumder A., Neamati N. and Ojwang JO. Inhibition of the human immunodeficiency virus type 1 integras by guanosine quartet structures. Biochemistry, 1996, 35:13762～13771
McCutchan F.E. HIV-1 genetic diversity. AIDS. 1996, 10:S13～S15
Meek T.D., Dayton B.D., Metcalf B.W., Dreyer G.B., Strickler J.E., Gorniak J.G., Rosenberg M., Moore M.L., Magaard V.W. and Debouck C. Human immunodeficiency virus 1 protease expressed in Escherichia coli behaves as a dimeric aspartic protease. Proceedings of National Acadcid Science of the USA, 1989, 86:1841～1845
Miles S.A. Protease inhibitors. International AIDS Society-USA. 1996, 47
Montagnier L., Gruest J., Chamaret S. and Dauguet S. A new human T-lymphotropic retrovirus: characterization and possible role in lymphadenopathy and acquired immune deficiency syndromes. In Gallo RC, Essex and Gross L, human T-cell leukemia/lymphoma virus. CoM Spring Harbor Laboratory, Cold Spring Harbor, N.Y. 1984
Montagnier L. Clavel F., Guyader., Guetard D., Salle M. and Alizon M. Molecular cloning and polymorphism of the human immune deficiency virus type 2. Nature, 1986, 324:691～695
Navia M.A., Fitzgerald P.M., McKeever B.M., Leu C.T., Hermbach J.C., Herber W.K., Sigal I.S., Darke P.L. and Springer J.P. Three-dimensional structure aspartyl protease from human immunodeficiency virus HIV-1. Nature, 1989, 337:615～620
Nie Z.L. Bergeron D., Subbramanian R.A, Yao X.J., Kougeau N. and Cohen E.A. The putative alpha helix 2 of human immunodeficiency virus type 1 Vpr contains a determinant which is responsible for the nuclear translocation of proviral DNA in growth arrested cells. Journal of Virology, 1998, 72:4104～4115
Ng T.B. and Huang B. Anti-human immunodeficiency vims(anit-HIV)naturai products with special emphasis on HIV reverse transcriptase inhibitors. Life Science. 1997, 61:933～949
Oude Essink B.B., Des A.T. and Berkhout B. HIV-1 reverse transcriptase discriminates against non-self tRNA primers. Journal of Molecular Biology, 1996, 264:243～254
Pearl L.H. and Taylor W.R. Sequence specificity of retroviral proteases. Nature, 1978, 328:482
Poon D.T.K., Li G. and aldovine A. Nucleocapsid and matrix protein contributions to selective Human immunodeficiency virus type 1 genomic RNA packaging. Journal of Virology, 1998, 72:1983～1993
Sagg M.S., Maseur H. and Micheal S. Strategies for continuing antiretroviral therapy. International AIDS Sonciety-USA, 1996, 4:11
Saitoh T, Amuro K. and Takaschi D. Antitumaor activity of Hypsizigus marmoreus Ⅱ Preventive effect against lung metastasis of lewis lung carcinoma. Yakugaku Zasshi, 1997, 117:1006
Schneider D.J., Feigon J., Hostomsky Z. and Gold L. High-affinity ssDNA inhibitor of the reverse transcriptase of type 1 human immunodeficiency virus. Biochemistry, 1995, 34:9599～9610
Spence R.A., Kati W.M., Anderson K.S. and Johnson K.A. Mechanisim of inhibition of H1V-1 reverse transcriptase by non-nucleoside inhibitors. Science, 1995, 267:988～993
Takechara M., Toyomasu T., Mori K. and Madanobu M. Isolation and antiviral activities of couble-stranded RNA from Lentinus edodes(shiitake). Kobe Journal of Medicinal Science, 1984, 30:25～34
Wang H.X. and Ng T.B. Examination of lectins, polysaccharopeptide, polysaccharide, alkaloid, coumarin and trypsin inhibitors for inhibitory activity against human immunodeficiency virus reverse transcriptase and glycohydrolases. Planta Medicine, 2001, 67:669～672
Walodawer A., Miller M., Jaskolski M., Sathyanarayana B.K., Baldwin E., Weber I.T., Selk L.M., Clawson L., Schnerder J. and Kent S.B. Conserver folding in retroviral proteases: crystal structure of a synthetin HIV-1 protease. Science, 1989, 245:616～621
Wyatt R. and Joseph S. The HIV-1 envelope glycoproteins: fusogens, antigens and immunogens. Science, 1998, 280:1884～1889
张兴权等．艾滋病毒感染与艾滋病．人民卫生出版社，1999，1～12
张惟杰．复合多糖生化研究技术．上海科学技术出版社，1987，209～215
金宁一．HIV基因结构及疫苗研究．中国病毒学，1997，12：285～296
张晓莉等．医学免疫和微生物学．军事医学科学出版社，1998，236～238
吴东儒．糖的生物化学．高等教育出版社．1987，621～623
侯云德．分子病毒学．学苑出版社．1990，480～488
肖盛元，郭顺星．国内真菌多糖结构研究现状．天然产物研究与开发，2002，12：81～86
阎隆飞，孙之荣．蛋白质分子结构．清华大学出版社，1999，27～29
孙崇荣，李玉民．蛋白质化学导论．复旦大学出版社，1991，68～70
黄年来．中国大型真菌原色图谱．中国农业出版社，1998，255～260
Anthony S. Fauci. The human immunodeficiency virus: infectivity and mechanisms of pathogenesis. Science, 1988, 239:617～622
Bagarazzi M.I., Boyer J.D. and Ayyavoo V. Nucleic acid-based vaccines as an approach to immunization against human immunodeficiency virus type-Ⅰ. Current Top Microbiological Immunology,1998, 226:107～143
Barreiro P., Soriana V. and Blanco F. Risks and benefits of replacing protease inhibitors by nevirapine in HIV-infected subjects under long-term successful triple combination therapy. AIDS, 2000, 14:807～812
Bartlett J.G. Protease inhibitor for HIV infection. Annals of Internal Medicine, 1996, 124: 1086～1087
Cai M., Huang Y., Caffrey M., Zheng R., Craigie R. and Clore GM. Solution structure of the N-terminal zinc binding domain of HIV integrase. Nature Structural Biology. 1997, 4:567～577
Chang R.S. and Yeung H.W. Inhibiton of growth of human immunodeficiency virus in vitro by crude extracts of Chinese medicinal herbs. Antiviral Research, 1988, 9:163～176
Chang S.T. and Miles P.G. Mushroom biology-A new discipline. The Mycologist, 1992, 6:64～65
Chihara G., Hamuro J., Maeda Y.Y., Ami Y. and Fukuoka F. Fraction and Purification of the polysaccharides with marked antitumor activity, especially Lentinan from Lentinus edodes. Cancer Research, 1970, 30:943～944
Chung R.S. Functional properties of edible mushroom. Nutrition Review, 1996, 54(11):91～94
Chopra K.F. and Tyring S.K. Current antiretroviral therapy in the treatment of HIV infection. Seminars Cutaneou Medical Surgery, 1997, 16:381～382
Collins R.A., Ng T.B., Fong W.P., Wan C.C. and Yeung H.W. A comparison of human immunodeficiency virus type Ⅰ infection by partially purified aqueous extracts of Chinese medicinal herbs. Life Sciences, 1997, 60:pc345～351
Cooper J.I. The nature of virus. In Virus and Environment(2nd ed.). London: Chapman & Hall, 1995, 1～33
Dalgeish A.G. and Beverly P.L. The CD4(T4) antigen is an essential compnent of the receptor of the AIDS retrovirus. Nature, 1984, 312:763～767
Debyser Z. HIV-1 Reverse transcription in viral DNA synthesis. Leuven: Leuven University Press. 1994, 23～24
Emernan M. and Malim M.H. HIV-1 regulatory/accessory genes: key to unraveling viral and host cell biology. Science, 1998, 280:1880～1884
Fesen M.R., Kohn K.W., Leteurtre F. and Pommier Y. Inhibitors of human immunodeficiency virus integrase. Proceedings of National Academy of Sciences of the USA, 1993, 90:2399～2340
Fisher R.J., Rein A., Fivashi M., Urbaneja M.A., Casas-finet J.R., Medaglia M. and Henderson L.E. Sequence-specific binding of human immunodeficiency virus type Ⅰ nucleacapsid protein to shoot oligonucleotides. Journal of Virology. 1998, 72:1902～1909
Francesco M.A., Caruso A., Fallacara F., Canaris A., Francesco D., Poiesi C., Licenziati S., Coalli M., Martinielli F., Fiorentini S. and Turano A. HIV p17 enhances lymphocyte proliferation and HIV-I replication after binding to a human serum factor. AIDS. 1998, 12:245～252
Gerton J.L., Sharron O., Mari O., Joseph D. and Patrick D. Effects of mutations in residues near the active site of human immunodeficiency virus type Ⅰ integrase on specific enzyme-substrate interaction. Journal of Virology. 1998, 72:5046～5055
Gotch F., Hardy G. and Imami N. Therapeutic vaccines in HIV-1 infection. Immunology Review, 1999,170:173～182
Goto T. and Komuro N. Projection structures of human immunodeficiency virus type 1(HIV-1) observed with high resolution electron microscopy. Journal of Electron Microscopy. 1994, 43:16～19
He J.L. CCR3 and CCR5 are co-receptors for HIV-1 infection of microgha. Nature. 1997, 385:645～649
Hammer S.M. Antiretroviral therapy: where are we? AIDS. 1998, 12:S181～S188
Hermann T., Meirer T., Gotte M. and Heumann H. The 'helix clamp' in HIV-1 reverse transeriptase nuleic acid binding motif common in nucleic acid polymerase. Nucleic Acid Research, 1994, 22:1686～1695
Hoffman T.L. Chemokines and co-receptors in HIV/SIV-host interaction. AIDS, 1998, S17～S26
Huang M.J. Incorporation of p160~(gag-pol) into virus particles requires the presence of both the major homology region and adjacent C-terminal capsid sequences with the Gag-Pol polyprotein. Journal of Virology. 1997, 4412～4418
Ikekawa T., Cehara N., Maeda Y., Nakanishi M. and Fukuoka E Antitumor activity of aqueous extracts of edible mushrooms. Cancer Research, 1969, 29:734～735
Jing N., Marchand C., Liu J., Mitra R., Hogan M.E. and Pommier Yves. Mechanism of inhibition of HIV-1 integrase by G-tetrad-forming oligonucleotides in vitro. Journal of Biological Chemistry. 2000, 275:21460～21467
Johnston M.I. HIV vaccines: problems and prospects. Hosp. Prac. (OffS. Ed.), 1997, 32:125～140
Kaneda T., Arai K. and Tokuda S. The effect of dried mushroom: Cortinellus shiitake on cholesterol metabolism in rats. Journal of Japanese Social Food Nutrition, 1964, 16:466～468
Khan E., Mack J.P., Katx R.A., Kullkosky J. and Skalka A.M. Retroviral integrase domains: DNA binding and the recognition of LTP sequences. Nucleic Acids Research, 1991, 19:581～860
Kraus E., James W. and Barclay A.N. Cutting Edge: noval RNA ligands able to bind CD4 antigen and inhibit CD4~+ T lymphocyte function. The journal of immunology. 1998, 160:5309～5212
Lafeuillade A. Effects of a comvination fo zidovudine, didanosine and lamivudine on primary human immunodeficiency virus type 1 infection. Journal of Infectious Disease. 1997,175～124
Levy J.A. HIV and the pathogenesis of AIDS. ASM Press, 1994
Littvak S. Retroviral Reverse Transcriptase. R. G. Lands Company, 1996
Luban J. Absconding with the chaperone: essential cyclophilin-gag interaction in HIV-1 virion. Cell. 1996, 87:1157～1159
Luciw P.A. Human immunodeficiency virus and their replication. In Field B.N. et al. ed Field Virology(3rd ed.). Philadelphia: Lippincott-Raven Publishers, 1996, 1881～1952
Mazumder A., Neamati N. and Ojwang JO. Inhibition of the human immunodeficiency virus type 1 integras by guanosine quartet structures. Biochemistry, 1996, 35:13762～13771
McCutchan F.E. HIV-1 genetic diversity. AIDS. 1996, 10:S13～S15
Meek T.D., Dayton B.D., Metcalf B.W., Dreyer G.B., Strickler J.E., Gorniak J.G., Rosenberg M., Moore M.L., Magaard V.W. and Debouck C. Human immunodeficiency virus 1 protease expressed in Escherichia coli behaves as a dimeric aspartic protease. Proceedings of National Acadcid Science of the USA, 1989, 86:1841～1845
Miles S.A. Protease inhibitors. International AIDS Society-USA. 1996, 47
Montagnier L., Gruest J., Chamaret S. and Dauguet S. A new human T-lymphotropic retrovirus: characterization and possible role in lymphadenopathy and acquired immune deficiency syndromes. In Gallo RC, Essex and Gross L, human T-cell leukemia/lymphoma virus. CoM Spring Harbor Laboratory, Cold Spring Harbor, N.Y. 1984
Montagnier L. Clavel F., Guyader., Guetard D., Salle M. and Alizon M. Molecular cloning and polymorphism of the human immune deficiency virus type 2. Nature, 1986, 324:691～695
Navia M.A., Fitzgerald P.M., McKeever B.M., Leu C.T., Hermbach J.C., Herber W.K., Sigal I.S., Darke P.L. and Springer J.P. Three-dimensional structure aspartyl protease from human immunodeficiency virus HIV-1. Nature, 1989, 337:615～620
Nie Z.L. Bergeron D., Subbramanian R.A, Yao X.J., Kougeau N. and Cohen E.A. The putative alpha helix 2 of human immunodeficiency virus type 1 Vpr contains a determinant which is responsible for the nuclear translocation of proviral DNA in growth arrested cells. Journal of Virology, 1998, 72:4104～4115
Ng T.B. and Huang B. Anti-human immunodeficiency vims(anit-HIV)naturai products with special emphasis on HIV reverse transcriptase inhibitors. Life Science. 1997, 61:933～949
Oude Essink B.B., Des A.T. and Berkhout B. HIV-1 reverse transcriptase discriminates against non-self tRNA primers. Journal of Molecular Biology, 1996, 264:243～254
Pearl L.H. and Taylor W.R. Sequence specificity of retroviral proteases. Nature, 1978, 328:482
Poon D.T.K., Li G. and aldovine A. Nucleocapsid and matrix protein contributions to selective Human immunodeficiency virus type 1 genomic RNA packaging. Journal of Virology, 1998, 72:1983～1993
Sagg M.S., Maseur H. and Micheal S. Strategies for continuing antiretroviral therapy. International AIDS Sonciety-USA, 1996, 4:11
Saitoh T, Amuro K. and Takaschi D. Antitumaor activity of Hypsizigus marmoreus Ⅱ Preventive effect against lung metastasis of lewis lung carcinoma. Yakugaku Zasshi, 1997, 117:1006
Schneider D.J., Feigon J., Hostomsky Z. and Gold L. High-affinity ssDNA inhibitor of the reverse transcriptase of type 1 human immunodeficiency virus. Biochemistry, 1995, 34:9599～9610
Spence R.A., Kati W.M., Anderson K.S. and Johnson K.A. Mechanisim of inhibition of H1V-1 reverse transcriptase by non-nucleoside inhibitors. Science, 1995, 267:988～993
Takechara M., Toyomasu T., Mori K. and Madanobu M. Isolation and antiviral activities of couble-stranded RNA from Lentinus edodes(shiitake). Kobe Journal of Medicinal Science, 1984, 30:25～34
Wang H.X. and Ng T.B. Examination of lectins, polysaccharopeptide, polysaccharide, alkaloid, coumarin and trypsin inhibitors for inhibitory activity against human immunodeficiency virus reverse transcriptase and glycohydrolases. Planta Medicine, 2001, 67:669～672
Walodawer A., Miller M., Jaskolski M., Sathyanarayana B.K., Baldwin E., Weber I.T., Selk L.M., Clawson L., Schnerder J. and Kent S.B. Conserver folding in retroviral proteases: crystal structure of a synthetin HIV-1 protease. Science, 1989, 245:616～621
Wyatt R. and Joseph S. The HIV-1 envelope glycoproteins: fusogens, antigens and immunogens. Science, 1998, 280:1884～1889
张兴权等．艾滋病毒感染与艾滋病．人民卫生出版社，1999，1～12
张惟杰．复合多糖生化研究技术．上海科学技术出版社，1987，209～215
金宁一．HIV基因结构及疫苗研究．中国病毒学，1997，12：285～296
张晓莉等．医学免疫和微生物学．军事医学科学出版社，1998，236～238
吴东儒．糖的生物化学．高等教育出版社．1987，621～623
侯云德．分子病毒学．学苑出版社．1990，480～488
肖盛元，郭顺星．国内真菌多糖结构研究现状．天然产物研究与开发，2002，12：81～86
阎隆飞，孙之荣．蛋白质分子结构．清华大学出版社，1999，27～29
孙崇荣，李玉民．蛋白质化学导论．复旦大学出版社，1991，68～70
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