Ababhali: Wang Xiaoyan, Zhao Eryu

Iyunithi: Isibhedlele saseJiaozhou, iSibhedlele saseDongfang esiManyaniswa neYunivesithi yaseTongji

Okwangoku, olona hlobo lwesampulu luphambili lokufunyaniswa kwe-pathogen ye-nucleic acid yokuphefumla kukuswabha komqala.Kukho iintlobo ezininzi zezisombululo zokugcinwa kwesampulu ezisetyenziswa ngokuqhelekileyo, zonke kufuneka zifakwe efrijini okanye zibe ngumkhenkce ukuze zithuthwe kwaye zigcinwe;Kunzima ukulawula yonke inkqubo yobushushu obuphantsi ngexesha lokuqokelela kunye nenkqubo yokuthutha, kwaye kunzima ukuqinisekisa ulawulo lomgangatho phambi kovavanyo lwesampulu.^[1-2].

I-RNase (i-RNase) yi-endonuclease eyenza i-hydrolyzes i-RNA, ngokukodwa ukuqhawula amabhondi e-phosphodiester phakathi kwee-nucleotides.I-molecule ye-RNase izinzile kakhulu, kukho iibhondi ze-disulfide kwisakhiwo, kwaye umsebenzi wayo awufuni ubukho be-divalent cations, ngoko ke i-RNase ayinakuguqulwa ngokulula, kwaye kulula ukuyibuyisela kwakhona emva kokushisa okuphezulu okanye ukusetyenziswa kwe-denaturants.Ii-RNases zohlulwe zibe zi-endogenous kunye nezingaphandle.Ii-endogenous RNases zinokukhutshwa ngaxeshanye xa iiseli zigqabhukile.Ke ngoko, ukuphelisa indima ye-endogenous RNases linyathelo elibaluleke kakhulu kwinkqubo yokutsalwa kwe-RNA.Exogenous RNases zisasazwe ngokubanzi.I-Rnases ikhona emoyeni, ulusu lomntu, iinwele kunye namathe, ezizizathu ezibalulekileyo zokuthotywa lula kwe-RNA.^[3].

Umbuzo wedatha

I-CANS-CL02-A009 "Ukusetyenziswa koMgangatho weLabhoratri yezoNyango kunye neziKhokelo zokuVunywa koBuchule kwiNdawo yokuxilongwa kweMolekyuli" kwiimfuno zobugcisa iphakamisa ukuba imigangatho efanelekileyo yomgangatho wamanzi kufuneka yenziwe ngokwesicelo;Izikhongozeli ezilahlwayo ezingangeni moya:

RNase/Ukuhlelwa

(1) RNase A

I-Ribonuclease A (i-RNase A), ephuma kwi-pancreas ye-bovine, i-endoribonuclease enokuhlasela ngokuthe ngqo i-3 'isiphelo se-pyrimidine i-residues kwi-RNA, ukusika i-cytosine okanye i-uracil eyenziwa yi-nucleotides esondeleyo.I-Phosphodiester bond, imveliso yokugqibela yokusabela yi-3 'pyrimidine nucleotide kunye ne-oligonucleotide ene-3' pyrimidine nucleotide ekupheleni.

(2) RNase T1

I-Ribonuclease T1 (i-RNase T1) ivela kwi-Aspergillus orjzae, isebenza ngokuthe ngqo kwi-3'-terminal phosphate ye-guanine, kwaye indawo yokuqhawula iphakathi kwe-3′ phosphate ye-guanine kunye ne-5′ hydroxyl ye-nucleotides ekufutshane.Imveliso yokugqibela yokusabela yi-3′guanylic acid kunye ne-oligonucleotide fragments kunye ne-3′guanylic acid ekugqibeleni.

(3) RNase H

I-Ribonuclease H (RNase H) yafunyanwa okokuqala kwinyama yethole i-thymus, kwaye i-encoding gene yayo iye yahlanganiswa kwi-Escherichia coli.Iyakwazi ukuthoba ngokuthe ngqo i-DNA: i-RNA strands kwi-RNA hybrid duplexes, ebangela ukuba i-oligonucleotides kunye ne-mononucleotides kunye ne-3'-OH kunye ne-5'-monophosphate ekupheleni, ayikwazi ukuthobisa i-DNA enye okanye i-double-stranded DNA okanye i-RNA.

RNase

Umsebenzi kunye nokusetyenziswa

I-Ribonuclease inokubangela ukuthotywa kwe-ribonucleic acid (RNA) kwaye inokwenziwa ngokufanelekileyo.I-ointment yonyango isetyenziselwa ukunyanga ukwenzakala kunye neentlungu ezidibeneyo.Ngokweengxelo, i-ribonuclease inokutshintsha i-metabolism ye-host cell, inhibit virus synthesis, inqanda ukwanda kwe-influenza virus in vitro, kwaye inqanda ukubunjwa kwe-vaccinia kunye ne-herpes virus kwi-embryos yenkukhu.Ukusetyenziswa kwekliniki ye-ribonuclease yemihla ngemihla ye-intramuscular injection ye-180 mg, iluncedo kunyango lwe-encephalitis ye-epidemic, i-ribonuclease inokubangela ukuthotywa kwe-ribonucleic acid (RNA), kwaye ngoku ingenziwa ngobuchule.I-ointment yonyango isetyenziselwa ukunyanga ukwenzakala kunye neentlungu ezidibeneyo.Ngokweengxelo, i-ribonuclease inokutshintsha i-metabolism ye-host cell, inhibit virus synthesis, inqanda ukwanda kwe-influenza virus in vitro, kwaye inqanda ukubunjwa kwe-vaccinia kunye ne-herpes virus kwi-embryos yenkukhu.Ukusetyenziswa kwekliniki ye-ribonuclease yemihla ngemihla ye-intramuscular injection ye-180 mg iluncedo kunyango lwe-encephalitis ye-epidemic.

RNase

Inhibitor Inkcazo

Inkcazo yeyunithi ye-inhibitor ye-RNase: Isixa se-enzyme efunekayo ukuvimbela i-50% yomsebenzi we-5ng RNase A yiyunithi enye.

Indlela i-RNase Inhibitors esebenza ngayo

I-Guanidine isothiocyanate:

I-Guanidine isothiocyanate yi-organic compound kunye ne-molecular formula C2H6N4S.Ikakhulu isetyenziselwa amayeza ebhayoloji, ii-reagents zekhemikhali, njl njl. I-Guanidine isothiocyanate yi-protein lysing agent, kwaye isoloko isetyenziswa njengeyona nxalenye iphambili yesisombululo se-lysis kwi-molecular diagnostic reagents.Iyakwazi ukukhupha izicubu, itshabalalise ubume beeseli kwaye ihlukanise i-nucleic acid kwi-nucleoprotein, kwaye ibe ne-denaturation enamandla kwi-RNase.Yeyona inhibitor isebenzayo ye-RNase okwangoku.

I-TRIzol iyinoveli iyonke i-reagent yokutsalwa kwe-RNA enokuthi ikhuphe ngokuthe ngqo i-RNA epheleleyo kwiiseli okanye izicubu.Iqulethe izinto ezifana ne-phenol kunye ne-guanidine isothiocyanate, enokuphazamisa ngokukhawuleza iiseli kunye nokuvimbela i-nuclease ekhutshwe ngamaseli.

(Nangona kunjalo, i-guanidine isothiocyanate iyingozi kwimpilo yabaphandi balebhu.)

RNasin:

I-Acid glycoprotein ekhutshwa kwisibindi sempuku okanye kwi-blastoderm yabantu.I-Rnasin yi-inhibitor engakhuphisaniyo ye-RNase, enokuthi ibophe kwii-RNases ezahlukeneyo ukuze ingasebenzi.

(Iinkcukacha ezingakumbi: https://www.foreivd.com/foreasy-rnase-inhibitor-product/)

Hiqondo lobushushu:

Ubushushu obuphezulu bukwayindlela eqhelekileyo yokuchithwa kweprotheyini.

I-Diethylpyrocarbonate (DEPC):

I-DEPC yinto eyomeleleyo kodwa engaphelelanga ye-RNase inhibitor, enokuthintela umsebenzi we-RNase ngokudibanisa ne-amino acid imidazole ring yeqela elisebenzayo le-RNase ukuya kwiiprotheni ze-denature.

I-Vanadyl ribonucleoside complex:

I-complex eyakhiwe yi-vanadium oxide ion kunye ne-nucleosides, ebophelela kwi-RNase ngendlela yezinto eziguqukayo, ezinokuthi zithintele ngokupheleleyo umsebenzi we-RNase.

enye:

I-SDS, i-urea, umhlaba we-diatomaceous, njl.

Uphononongo lweNgcali

Li Yujie Chief Technician

UMlawuli weSebe leLebhu, iSibhedlele saseJiaozhou, iSibhedlele saseDongfang esidityaniswe neYunivesithi yaseTongji

Ukuze kuthintelwe ukubola kwe-RNase yangaphandle, iimaski, iiglavu kunye neminqwazi kufuneka zinxitywe kwaye zitshintshwe rhoqo ngexesha lenkqubo yokutsalwa kwe-RNA.Zonke izixhobo zeglasi kufuneka zibhakwe kwi-oven yokumisa kwi-200 ° C ngaphezu kweeyure ezingama-2.Izinto ezisetyenziselwa ukubhaka, njengeplastiki, kufuneka ziphathwe ngamanzi e-DEPC, emva koko zihlanjwe ngamanzi adibeneyo.Iirejenti okanye izixhobo ezisetyenziselwa ukukhutshwa, ukugcinwa kunye nokuchongwa kwe-RNA kufuneka zinikezelwe kwi-RNA, kwaye indawo yokusebenza ye-RNA ezimeleyo kufuneka imiselwe.

iimbekiselo:

[1] Smith-Vaughan HC, Binks MJ, Beissbarth J, et al.Ibhaktheriya kunye neentsholongwane kwi-nasopharynx ngokukhawuleza ngaphambi kokuqala kosulelo olusezantsi lokuphefumla kubantwana base-Australia [J].Eur J Clin Microbiol Infect Dis,2018,37 (9): 1785-1794.

[2] Isebe loLawulo lweSibhedlele lweSibhedlele soMbutho waseTshayina woKhuseleko lwezoNyango.Izikhokelo zokuqokelela kunye nokuhlolwa kweesampuli ze-microbial kliniki [J].Ijenali yaseTshayina yoSulelo lweSibhedlele, i-2018 (20): 3192-3200.

[3] “Uvavanyo lweKlinikhi ngamawaka alishumi Kutheni uVavanyo lweBhayoloji yeMolekyuli”