Ubuhlungu benyoka buqukethe ama-enzyme enza i-hydrolyze i-carboxyl ester bond.Ama-substrates e-hydrolysis yi-phospholipids, i-acetylcholine ne-acetate enamakha.Lawa ma-enzyme afaka izinhlobo ezintathu: i-phospholipase, i-acetylcholinesterase ne-esterase enamakha.I-arginine esterase ebuhlungwini benyoka ingaphinda ikhiqize i-arginine yokwenziwa noma i-lysine, kodwa ikakhulukazi i-hydrolyze amabhondi e-protein peptide ngokwemvelo, ngakho ingeye-protease.Ama-enzyme okuxoxwe ngawo lapha asebenza kuphela kuma-ester substrates futhi awakwazi ukusebenza kunoma iyiphi ibhondi ye-peptide.Phakathi kwalawa ma-enzyme, imisebenzi yebhayoloji ye-acetylcholinesterase ne-phospholipase ibaluleke kakhulu futhi ifundwe ngokugcwele.Obunye ubuthi bezinyoka bunomsebenzi oqinile we-esterase onuka kamnandi, ongenza i-hydrolyze i-p-nitrophenyl ethyl ester, i-a – noma i-P-naphthalene acetate ne-indole ethyl ester.Namanje akwaziwa ukuthi lo msebenzi ukhiqizwa i-enzyme ezimele noma umphumela owaziwayo we-carboxylesterase, ingasaphathwa eyokubaluleka kwayo kwezinto eziphilayo.Lapho ubuthi be-Agkistrodon halys Japonicus buphendulwa nge-p-nitrophenyl ethyl ester ne-indole ethyl ester, ama-hydrolysates e-p-nitrophenol ne-indole phenol awatholakalanga;Ngokuphambene nalokho, uma lawa ma-esters esabela nge-cobra Zhoushan subspecies i-snake poison and Bungarus multicinctus snake poison, azokwenziwa i-hydrolyzed ngokushesha.Kuyaziwa ukuthi lezi zihlungu ze-cobra zinomsebenzi oqinile we-cholinesterase, ongase ube nesibopho se-hydrolysis yama-substrates angenhla.Eqinisweni, uMclean et al.(1971) ibike ukuthi ubuthi benyoka abaningi bomndeni weCobra bungakhiqiza i-hydrolyze indole ethyl ester, naphthalene ethyl ester kanye ne-butyl naphthalene ester.Lobu buhlungu benyoka buphuma kubo: imfezi, imfezi emnyama nentamo emnyama, imfezi enamalebe amnyama, imfezi egolide, imfezi yaseGibhithe, i-king cobra, i-golden cobra mamba, i-black mamba ne-white lipped mamba (D. aw usayazi i-eastern rhombola rattlesnake
Ubuhlungu benyoka bungakhiqiza i-hydrolyze i-methyl indole ethyl ester, okuyi-substrate yokunquma umsebenzi we-cholinesterase ku-serum, kodwa lobu buthi benyoka abubonisi umsebenzi we-cholinesterase.Lokhu kukhombisa ukuthi kune-esterase engaziwa ebuhlungwini be-cobra, obuhlukile kune-cholinesterase.Ukuze uqonde uhlobo lwale enzyme, kudingeka omunye umsebenzi wokuhlukanisa.
1, Phospholipase A2
(I) Uhlolojikelele
I-Phospholipase iyi-enzyme ekwazi ukwenza i-hydrolyze i-glyceryl phosphate.Kunezinhlobo ezi-5 ze-phospholipase emvelweni, okuyi-phospholipase A2 ne-phospholipase
A. , i-phospholipase B, i-phospholipase C ne-phospholipase D. Ubuthi benyoka ikakhulukazi buqukethe i-phospholipase A2 (PLA2), ubuthi obumbalwa bezinyoka buqukethe i-phospholipase B, namanye ama-phospholipase atholakala kakhulu ezicutshini zezilwane nakumabhaktheriya.Umdwebo 3-11-4 ubonisa indawo yesenzo yalezi phospholipases ku-substrate hydrolysis.
Phakathi kwama-phospholipases, i-PLA2 ifundwe kabanzi.Kungase kube i-enzyme efundwe kakhulu ebuhlungwini benyoka.I-substrate yayo iyisibopho se-ester endaweni yesibili ye-Sn-3-glycerophosphate.Le enzyme itholakala kakhulu ebuhlungwini bezinyoka, ubuthi bezinyosi, isihlungu sikafezela kanye nezicubu zezilwane, kanti i-PLA2 igcwele ezinhlamvini ezine zezinyoka zomndeni.Ngenxa yokuthi le enzyme iphula amangqamuzana egazi abomvu futhi ibangele i-hemolysis, ibizwa nangokuthi "i-hemolysin".Abanye abantu baphinde babize i-PLA2 i-hemolytic lecithinase.
U-Ludeeke waqala wathola ukuthi ubuthi benyoka bungakhiqiza isakhi se-hemolytic ngokusebenza ku-lecithin ngama-enzyme.Kamuva, u-Delezenne et al.kwafakazela ukuthi lapho ubuthi be-cobra busebenza ku-serum yamahhashi noma i-yolk, yakha into eyi-hemolytic.Manje sekuyaziwa ukuthi i-PLA2 ingakwazi ukusebenza ngokuqondile ku-phospholipids ye-membrane ye-erythrocyte, ibhubhise ukwakheka kwe-erythrocyte membrane futhi ibangele i-hemolysis eqondile;Ingase futhi isebenze ku-serum noma i-lecithin eyengeziwe ukuze ikhiqize i-hemolytic lecithin, esebenza kumaseli abomvu egazi ukukhiqiza i-hemolysis engaqondile.Nakuba i-PLA2 igcwele emindenini emine yobuthi bezinyoka, okuqukethwe kwama-enzyme kubuthi benyoka obuhlukahlukene kuhluke kancane.I-Rattlesnake (C
Ubuhlungu benyoka bubonise kuphela umsebenzi obuthakathaka we-PLA2.Ithebula 3-11-11 libonisa ukuqhathaniswa komsebenzi we-PLA2 wobuthi obukhulu obuyi-10 bezinyoka ezinobuthi e-China.
Ithebula 3-11-11 Ukuqhathaniswa kwemisebenzi ye-phospholipase VIII yezinhlayiya eziyi-10 zenyoka eChina
Ubuhlungu benyoka
Ukukhululwa kwamafutha
I-aliphatic acid,
Cjumol/mg)
Umsebenzi we-Hemolytic CHU50/^ g * ml)
Ubuhlungu benyoka
Khipha ama-fatty acids
(^raol/mg)
Umsebenzi we-Hemolytic “(HU50/ftg * 1111)
Najaja atra
9. 62
ishumi nanye
I-Micracephal ophis
amaphuzu amahlanu uziro oyedwa
ama-kalyspallas
8. 68
izinkulungwane ezimbili namakhulu ayisishiyagalombili
gracilis
V, i-actus
7. 56
* * #
Ofiyophagus Hana
amaphuzu amathathu ayisishiyagalombili amabili
ikhulu namashumi amane
I-Bnugarus fasctus
7,56
amakhulu amabili namashumi ayisishiyagalombili
B. multicinctus
iphuzu elilodwa isishiyagalolunye nesithupha
amakhulu amabili namashumi ayisishiyagalombili
I-Viper ne-russelli
amaphuzu ayisikhombisa uziro amathathu
T, i-mucrosquamatus
iphuzu elilodwa isishiyagalombili nesihlanu
I-Siamensis
T. stejnegeri
0.97
(2) Ukuhlukaniswa nokuhlanzwa
Okuqukethwe kwe-PLA2 ebuhlungwini benyoka kukhulu, futhi kuzinzile ekushiseni, i-asidi, i-alkali ne-denaturant, ukuze kube lula ukuyihlanza nokuhlukanisa i-PLA2.Indlela evamile iwukuba uqale ukuhlunga ijeli ebuhlungwini obungahluziwe, bese wenza i-ion exchange chromatography, futhi isinyathelo esilandelayo singaphindwa.Kufanele kuqashelwe ukuthi ukumisa iqhwa kwe-PLA2 ngemva kwechromatography ye-ion-exchange akumele kubangele ukuhlanganisa, ngoba inqubo yokumisa iqhwa ngokuvamile inyusa amandla e-ionic ohlelweni, okuyisici esibalulekile esibangela ukuhlanganisa kwe-PLA2.Ngaphezu kwalezi zindlela ezijwayelekile ezingenhla, lezi zindlela ezilandelayo nazo sezamukelwe: ① Wells et al.② I-analogue engaphansi ye-PLA2 isetshenziswe njenge-ligand ye-affinity chromatography.Le ligand ingabopha ku-PLA2 ebuhlungwini benyoka nge-Ca2+.I-EDTA isetshenziswa kakhulu njenge-elient.Ngemva kokususwa kwe-Ca2+, ukuhlobana phakathi kwe-PLA2 ne-ligand kuncipha, futhi kungahlukaniswa ne-ligand.Abanye basebenzisa i-30% ye-organic solution noma i-6mol/L urea njengendlela engaqondakali.③ I-Hydrophobic chromatography yenziwe nge-PheiiylSephar0SeCL-4B ukuze kukhishwe umkhondo we-PLA2 ku-cardiotoxin.④ I-anti PLA2 antibody isetshenziswe njenge-ligand ukwenza i-affinity chromatography ku-PLA2.
Kuze kube manje, inani elikhulu lobuthi benyoka PLAZ selihlanziwe.Thu et al.(1977) kufakwe kuhlu lwe-PLA2 ehlanjululwe ebuhlungwini benyoka ngaphambi kuka-1975. Eminyakeni yamuva, inani elikhulu lama-athikili mayelana nokuhlukaniswa nokuhlanzwa kwe-PLA2 liye labikwa njalo ngonyaka.Lapha, sigxile ekuhlukaniseni nasekuhlanzweni kwe-PLA yizazi zaseShayina.
U-Chen Yuancong et al.(1981) yahlukanisa izinhlobo ezintathu ze-PLA2 ebuhlungwini be-Agkistrodon halys Pallas e-Zhejiang, engahlukaniswa ibe i-acidic, neutral kanye ne-alkaline i-PLA2 ngokuya ngamaphuzu azo ahlukene kagesi.Ngokobuthi bayo, i-PLA2 engathathi hlangothi inobuthi kakhulu, ekhonjwe njenge-presynaptic neurotoxin Agkistrodotoxin.I-alkaline PLA2 ayinabo ubuthi obuncane, futhi i-PLA2 ene-asidi cishe ayinabo ubuthi.Wu Xiangfu et al.(I-1984) iqhathanise izici zama-PLA2 amathathu, okuhlanganisa isisindo samangqamuzana, ukwakheka kwe-amino acid, i-N-terminal, iphuzu le-isoelectric, ukuzinza okushisayo, umsebenzi we-enzyme, ubuthi kanye nomsebenzi we-hemolytic.Imiphumela yabonisa ukuthi babenesisindo esifanayo samangqamuzana nokuzinza kokushisa, kodwa babe nomehluko omkhulu kwezinye izici.Esicini somsebenzi we-enzyme, umsebenzi we-enzyme ye-asidi wawuphakeme kunomsebenzi we-alkaline enzyme;Umthelela we-hemolytic we-alkaline enzyme kumagundane egazi abomvu wawunamandla, ulandelwa yi-enzayimu engathathi hlangothi, kanye ne-enzayimu ye-asidi icishe ibe ne-hemolyzed.Ngakho-ke, kucatshangwa ukuthi umphumela we-hemolytic we-PLAZ uhlobene nokukhokhiswa kwe-molecule ye-PLA2.UZhang Jingkang et al.(1981) benze amakristalu e-Agkistrodotoxin.UTu Guangliang et al.(1983) ibike ukuthi i-PLA enobuthi enephuzu le-isoelectric le-7. 6 yahlukaniswa futhi yahlanzwa ebuhlungwini be-Vipera rotundus obuvela eFujian, kanye nezakhiwo zayo ezingokomzimba namakhemikhali, ukwakheka kwe-amino acid kanye nokulandelana kwezinsalela ze-amino acid ezingama-22 ku-N. -ukuphela kwanqunywa.U-Li Yuesheng et al.(1985) yahlukanisa futhi yahlanza enye i-PLA2 ebuhlungwini beViper rotundus eFujian.I-subunit ye-PLA2 * ingu-13 800, iphoyinti le-isoelectric ngu-10.4, futhi umsebenzi othize ngu-35/xnioI/miri mg. Nge-lecithin njenge-substrate, i-pH elungile ye-enzyme ingu-8.0 futhi izinga lokushisa elilungile lingu-65 ° C. I-LD5 ijovwe ngomthambo wamagundane.Kungu-0.5 ± 0.12mg/kg.Le enzyme inemiphumela esobala ye-anticoagulant ne-hemolytic.I-molecule ye-PLA2 enobuthi iqukethe izinsalela ezingu-123 zezinhlobo ezingu-18 zama-amino acid.I-molecule inothile nge-cysteine (14), i-aspartic acid (14) ne-glycine (12), kodwa iqukethe i-methionine eyodwa kuphela, futhi i-N-terminal yayo iyinsalela ye-serine.Uma kuqhathaniswa ne-PLA2 ehlukaniswe i-Tuguang, isisindo samangqamuzana kanye nenani lezinsalela ze-amino acid zama-isoenzymes amabili afana kakhulu, futhi ukwakheka kwe-amino acid nakho kufana kakhulu, kodwa inani le-aspartic acid nezinsalela ze-proline lihlukile.I-Guangxi king cobra snake poison iqukethe i-PLA2 ecebile.Shu Yuyan et al.(1989) ihlukanise i-PLA2 ebuhlungwini, enomsebenzi othize ophindwe izikhathi ezingu-3.6 kunobuthi bokuqala, isisindo semolekyuli esingu-13000, ukwakheka kwezinsalela ze-amino acid eziyi-122, iphoyinti le-isoelectric elingu-8.9, kanye nokuzinza okuhle kwe-thermal.Ngokubona nge-electron microscope umphumela we-PLA2 eyisisekelo kumaseli abomvu egazi, kuyabonakala ukuthi inomthelela osobala kulwelwesi lwamangqamuzana egazi abomvu, kodwa alinawo umthelela osobala kumangqamuzana egazi abomvu ezimbuzi.Le PLA2 inomphumela osobala wokuvinjelwa kwejubane le-electrophoretic lamaseli abomvu egazi kubantu, ezimbuzini, onogwaja kanye nezingulube ze-Guinea.U-Chen et al.Le enzyme ingavimbela ukuhlanganiswa kweplatelet okubangelwa i-ADP, i-collagen ne-sodium arachidonic acid.Uma ukugxilisa kwe-PLA2 kungu-10/xg/ml~lOOjug/ml, ukuhlanganiswa kweplatelet kuvinjelwa ngokuphelele.Uma ama-platelet awashiwe asetshenziswa njengezinto zokwakha, i-PLA2 ayikwazanga ukuvimbela ukuhlanganisa ekugxilweni kuka-20Mg/ml.I-Aspirin iyi-inhibitor ye-cyclooxygenase, engavimbela umphumela we-PLA2 kuma-platelet.I-PLA2 ingase ivimbele ukuhlangana kweplatelet nge-hydrolyzing arachidonic acid ukuze ihlanganise i-thromboxane A2.Ukwakheka kwesixazululo se-PLA2 okukhiqizwe yi-Agkistrodon halys Pallas venom esifundazweni sase-Zhejiang kwacwaningwa kusetshenziswa i-dichroism eyindilinga, i-fluorescence kanye nokumuncwa kwe-UV.Imiphumela yokuhlola yabonisa ukuthi ukwakheka kweketango eliyinhloko lale enzyme kwakufana nohlobo olufanayo lwe-enzyme oluvela kwezinye izinhlobo zezilwane nesizukulwane, ukwakheka kwamathambo kwakunokumelana okuhle nokushisa, futhi uguquko lwesakhiwo endaweni ye-asidi lwalubuyiseleka emuva.Inhlanganisela ye-activator Ca2+ kanye ne-enzyme ayithinti imvelo yezinsalela ze-tryptophan, kuyilapho i-inhibitor Zn2+ yenza okuphambene.Indlela inani le-pH lesisombululo elithinta ngayo umsebenzi we-enzyme ihlukile kuma-reagents angenhla.
Enqubweni ye-PLA2 yokuhlanzwa kwesihlungu senyoka, into esobala ukuthi ubuthi benyoka buqukethe iziqongo ezimbili noma ngaphezulu ze-PLA2.Lesi simo singachazwa kanje: ① ngenxa yokuba khona kwama-isozyme;② Olunye uhlobo lwe-PLA2 lwenziwe i-polymerized lube izingxube ezihlukahlukene ze-PLA2 ezinesisindo samangqamuzana ahlukahlukene, iningi lazo elikububanzi obuyi-9 000~40 000;③ Inhlanganisela ye-PLA2 nezinye izingxenye zesihlungu senyoka yenza i-PLA2 ibe nzima;④ Ngenxa yokuthi ibhondi ye-amide ku-PLA2 yenziwe nge-hydrolyzed, inkokhiso iyashintsha.① Futhi ② zivamile, ngaphandle nje okumbalwa, njenge-PLA2 ku-CrWa/w ubuthi benyoka
Kunezimo ezimbili: ① kanye ②.Isimo sesithathu sitholakale ku-PLA2 ebuhlungwini bezinyoka ezilandelayo: i-Oxyranus scutellatus, i-Parademansia microlepidota, i-Bothrops a ^>er, i-Palestinian viper, i-sand viper, ne-rattlesnake esabekayo km.
Umphumela wecala ④ wenza isivinini sokufuduka se-PLA2 sishintshe ngesikhathi se-electrophoresis, kodwa ukwakheka kwe-amino acid akushintshi.Ama-peptide angaphulwa nge-hydrolysis, kodwa ngokuvamile asaboshelwe ndawonye ngamabhondi e-disulfide.Ubuthi be-eastern pit rattlesnake buqukethe izinhlobo ezimbili ze-PLA2, ezibizwa ngokuthi uhlobo a kanye nohlobo lwe-p PLA2 ngokulandelanayo.Umehluko phakathi kwalezi zinhlobo ezimbili ze-PLA2 yi-amino acid eyodwa kuphela, okungukuthi, i-glutamine ku-molecule eyodwa ye-PLA2 ithathelwa indawo yi-glutamic acid kwenye i-molecule ye-PLA2.Nakuba isizathu esiqondile salo mehluko singacacile, ngokuvamile kukholakala ukuthi sihlobene nokuchithwa kwe-PLA2.Uma i-PLA2 ebuhlungwini benyoka yasePalestine igcinwa ifudumele ngobuthi obungahluziwe, amaqembu okugcina kuma-molecule ayo e-enzyme azoba maningi kunangaphambili.Kusuka ku-C PLA2 ehlukanisiwe nobuthi benyoka ine-N-terminal emibili ehlukene, futhi isisindo sayo semolekyuli singu-30000. Lesi simo singase sibangwe i-asymmetric dimer ye-PLA2, efana ne-symmetric dimer eyakhiwe i-PLA2 ebuhlungwini be-eastern diamondback rattlesnake. kanye ne-western diamondback rattlesnake.Imfezi yase-Asia yakhiwe izinhlobonhlobo eziningi, ezinye zazo ezingacacile kahle ngokwezigaba.Isibonelo, lokho okwakubizwa ngokuthi i-Cobra Outer Caspian subspecies manje sekuyaziwa
Kufanele kubhekwe i-Outer Caspian Sea Cobra.Njengoba kunezinhlobo eziningi ze-subspecies futhi zixutshwe ndawonye, ukwakheka kwesihlungu senyoka kuyahluka kakhulu ngenxa yemithombo ehlukene, futhi okuqukethwe kwe-PLA2 isozymes nakho kuphezulu.Isibonelo, ubuthi be-cobra
Okungenani izinhlobo eziyi-9 ze-PLA2 isozymes zezinhlobo ze-r ^ll zatholakala kuzo, futhi izinhlobo eziyisi-7 zama-isozymes e-PLA2 zitholwe ebuhlungwini bezinhlobo ze-cobra Caspian.Durkin et al.(1981) itadishe okuqukethwe kwe-PLA2 kanye nenani lama-isozyme ebuhlungwini benyoka obuhlukene, okuhlanganisa ubuthi be-cobra obungu-18, ubuthi be-mamba obu-3, ubuthi be-viper obu-5, ubuthi benyoka ye-rattlesnake obu-3 kanye nobuthi benyoka yasolwandle obu-3.Ngokuvamile, umsebenzi we-PLA2 we-cobra venom uphezulu, nama-isozyme amaningi.Umsebenzi we-PLA2 kanye nama-isozymes we-viper venom aphakathi.Umsebenzi we-PLA2 wobuthi be-mamba kanye nobuthi be-rattlesnake uphansi kakhulu noma awukho umsebenzi we-PLA2.Umsebenzi we-PLA2 wobuthi benyoka yasolwandle nawo uphansi.
Eminyakeni yamuva nje, akuzange kubikwe ukuthi i-PLA2 ebuhlungwini benyoka ikhona ngendlela ye-dimer esebenzayo, njenge-eastern rhombophora rattlesnake (C. snake venom iqukethe uhlobo a kanye nohlobo P PLA2, kokubili okuhlanganisa amayunithi amabili afanayo. , futhi i-dimerase kuphela enayo
Umsebenzi.Shen et al.futhi uphakamise ukuthi i-dimer ye-PLA2 kuphela yobuhlungu benyoka iwuhlobo olusebenzayo lwe-enzyme.Ucwaningo lwesakhiwo sendawo futhi kufakazela ukuthi i-PLA2 ye-western diamondback rattlesnake ikhona ngendlela ye-dimer.Inhlanganisela ye-Piscivorous
Kunezinhlobo ezimbili ze-PLA ^ Ei kanye ne-E2 yobuthi benyoka, lapho i-仏 ikhona ngesimo se-dimer, i-dimer iyasebenza, futhi i-monomer yayo ehlukanisiwe ayisebenzi.U-Lu Yinghua et al.(1980) waqhubeka wafunda izici zomzimba namakhemikhali kanye ne-reaction kinetics ka-E. Jayanthi et al.(1989) yahlukanisa i-PLA2 eyisisekelo (VRVPL-V) ebuhlungwini bebululu.Isisindo samangqamuzana e-PLA2 ye-monomer yi-10000, enemiphumela ebulalayo, i-anticoagulant kanye ne-edema.I-enzyme ingakwazi ukupholisa ama-polymers anesisindo samangqamuzana ahlukene ngaphansi kwesimo se-PH 4.8, futhi izinga le-polymerization nesisindo samangqamuzana ama-polymers akhuphuka ngokunyuka kwezinga lokushisa.Isisindo samangqamuzana e-polymer ekhiqizwa ku-96 ° C singu-53 100, futhi umsebenzi we-PLA2 wale polymer ukhuphuka ngamabili.
Isikhathi sokuthumela: Nov-18-2022