Ama-Peptides ayikilasi lezinhlanganisela ezakhiwe ngokuxhunywa kwama-amino acid amaningi ngamabhondi e-peptide.Zitholakala yonke indawo ezintweni eziphilayo.Kuze kube manje, amashumi ezinkulungwane zama-peptide atholakele ezintweni eziphilayo.Ama-Peptide adlala indima ebalulekile ekulawuleni imisebenzi esebenzayo yezinhlelo ezihlukahlukene, izitho, izicubu namaseli kanye nemisebenzi yokuphila, futhi avame ukusetshenziswa ekuhlaziyeni ukusebenza, ucwaningo lwama-antibody, ukuthuthukiswa kwezidakamizwa neminye imikhakha.Ngokuthuthukiswa kwe-biotechnology kanye nobuchwepheshe be-peptide synthesis, imithi eminingi ye-peptide iye yathuthukiswa futhi yasetshenziswa emtholampilo.
Kunezinhlobonhlobo eziningi zokuguqulwa kwe-peptide, ezingahle zihlukaniswe zibe ukuguqulwa kokuthunyelwe kanye nokuguqulwa kwenqubo (kusetshenziswa ukuguqulwa kwe-amino acid etholakala), nokuguqulwa kwe-N-terminal, ukuguqulwa kwe-C-terminal, ukuguqulwa kwe-side chain, ukuguqulwa kwe-amino acid, ukuguqulwa kwamathambo, njll., kuye ngesayithi lokuguqulwa (Umfanekiso 1).Njengendlela ebalulekile yokuguqula ukwakheka kweketango eliyinhloko noma amaqembu amaketanga aseceleni amaketanga e-peptide, ukuguqulwa kwe-peptide kungashintsha ngokuphumelelayo izakhiwo ezingokomzimba nezamakhemikhali zezinhlanganisela ze-peptide, kukhuphule ukuncibilika kwamanzi, kunwebe isikhathi sesenzo ku-vivo, kuguqule ukusatshalaliswa kwawo kwezinto eziphilayo, kuqede i-immunogenicity. , nciphisa imiphumela emibi enobuthi, njll. Kuleli phepha, amasu amaningana amakhulu okuguqula i-peptide nezici zawo ziyethulwa.
1. Ukuhamba ngebhayisikili
Ama-cyclic peptide anezinhlelo eziningi zokusebenza ku-biomedicine, futhi ama-peptide amaningi emvelo anomsebenzi webhayoloji angama-cyclic peptides.Ngenxa yokuthi ama-peptide angama-cyclic avame ukuqina kakhulu kune-linear peptide, amelana ngokwedlulele ohlelweni lokugaya ukudla, angaphila emgudwini wokugaya ukudla, futhi abonise ukuhlobana okuqinile kwama-receptors okuqondiwe.Ukuhamba ngebhayisikili kuyindlela eqondile kakhulu yokwenza ama-peptide e-cyclic, ikakhulukazi ama-peptide anamathambo amakhulu esakhiwo.Ngokwemodi ye-cyclization, ingahlukaniswa ngohlobo lwe-chain-side chain, uhlobo lwe-terminal - side chain, uhlobo lwetheminali - uhlobo lwetheminali (uhlobo lokugcina ukuya ekugcineni).
(1) i-sidechain-to-sidechain
Uhlobo oluvame kakhulu lwe-side-chain to side-chain cyclization ibhuloho le-disulfide phakathi kwezinsalela ze-cysteine.Le cyclization yethulwa yizinsalela ezimbili ze-cysteine ezivinjiwe bese zifakwa i-oxid ukuze zakhe amabhondi e-disulfide.I-Polycyclic synthesis ingafinyelelwa ngokususa okukhethiwe kwamaqembu okuvikela i-sulfhydryl.Ukuhamba ngebhayisikili kungenziwa ku-solvent yangemuva kokuhlukana noma ku-pre-dissociation resin.Ukuhamba ngebhayisikili kuma-resin kungase kungasebenzi kahle kunokusebenzisa i-solvent cyclization ngoba ama-peptide kuma-resin awakhiqizi ama-cyclified conformations.Olunye uhlobo lwe-side-chain - side chain cyclization ukwakheka kwesakhiwo se-amide phakathi kwe-aspartic acid noma izinsalela ze-glutamic acid kanye nesisekelo se-amino acid, okudinga ukuthi iqembu lokuvikela iketango elisemaceleni likwazi ukususwa ngokukhetha ku-polypeptide noma ku-resin noma ngemuva kokuhlukaniswa.Uhlobo lwesithathu lwe-side-chain - i-side chain cyclization ukwakheka kwe-diphenyl ether nge-tyrosine noma i-p-hydroxyphenylglycine.Lolu hlobo lokuhamba ngebhayisikili emikhiqizweni yemvelo lutholakala kuphela emikhiqizweni yamagciwane, futhi imikhiqizo yamabhayisikili ngokuvamile inevelu yokwelapha engaba khona.Ukulungiswa kwalezi zinhlanganisela kudinga izimo zokusabela eziyingqayizivele, ngakho-ke azivamile ukusetshenziswa ekuhlanganiseni ama-peptide avamile.
(2) i-terminal-to-sidechain
I-Terminal-side cycle cyclization ngokuvamile ihlanganisa i-C-terminal neqembu le-amino le-lysine noma i-ornithine side chain, noma i-N-terminal ene-aspartic acid noma iketango laseceleni le-glutamic acid.Okunye ukujikeleza kwe-polypeptide kwenziwa ngokwakha amabhondi e-ether phakathi kwetheminali C kanye namaketango aseceleni e-serine noma e-threonine.
(3) Uhlobo lwetheminali noma lwekhanda nomsila
Ama-polypeptides e-Chain angenziwa ngebhayisikili ku-solvent noma amiswe ku-resin by side chain cyclation.Ukugxila okuphansi kwama-peptide kufanele kusetshenziswe ku-solvent centralization ukugwema i-oligomerization yama-peptide.Isivuno se-head-to-tail synthetic ring polypeptide sincike ekulandelaneni kwe-polypeptide yeketango.Ngakho-ke, ngaphambi kokulungiselela ama-peptide angama-cyclic ngezinga elikhulu, umtapo wezincwadi ongase ube nama-peptide omthofu oboshiwe kufanele uqale udalwe, ulandelwe wukuhamba ngamabhayisikili ukuze kutholwe ukulandelana okunemiphumela engcono kakhulu.
2. I-N-methylation
I-N-methylation ekuqaleni ivela kuma-peptide emvelo futhi yethulwa ekuhlanganiseni kwe-peptide ukuvimbela ukwakheka kwamabhondi e-hydrogen, ngaleyo ndlela yenza ama-peptide amelane kakhulu nokuwohloka kwezinto eziphilayo kanye nokuvunyelwa.Ukuhlanganiswa kwama-peptide kusetshenziswa okuphuma kokuphuma kwe-N-methylated amino acid kuyindlela ebaluleke kakhulu.Ngaphezu kwalokho, ukusabela kwe-Mitsunobu kwe-N-(2-nitrobenzene sulfonyl chloride) i-polypeptide-resin intermediate ene-methanol ingasetshenziswa.Le ndlela isetshenziselwe ukulungisa imitapo yolwazi ye-cyclic peptide equkethe i-N-methylated amino acid.
3. Phosphorylation
I-Phosphorylation ingenye yezinguquko ezivame kakhulu ngemuva kokuhumusha emvelweni.Emangqamuzaneni omuntu, ngaphezu kwama-30% amaprotheni ane-phosphorylated.I-phosphorylation, ikakhulukazi i-phosphorylation ebuyela emuva, idlala indima ebalulekile ekulawuleni izinqubo eziningi zamaselula, njengokudluliswa kwesignali, ukubonakaliswa kofuzo, umjikelezo wamaseli kanye nokulawulwa kwe-cytoskeleton, kanye ne-apoptosis.
I-phosphorylation ingabonwa ezinhlobonhlobo zezinsalela ze-amino acid, kodwa okuhlosiwe okuvame kakhulu kwe-phosphorylation yizinsalela ze-serine, i-threonine, ne-tyrosine.I-Phosphotyrosine, i-phosphothreonine, kanye ne-phosphoserine derivatives ingafakwa kuma-peptide ngesikhathi sokuhlanganiswa noma yakhelwe ngemva kokuhlanganiswa kwe-peptide.I-phosphorylation ekhethiwe ingafinyelelwa kusetshenziswa izinsalela ze-serine, i-threonine, ne-tyrosine esusa amaqembu okuzivikela ngokukhetha.Amanye ama-reagents e-phosphorylation angakwazi futhi ukwethula amaqembu e-phosphoric acid ku-polypeptide ngokuguqulwa kweposi.Eminyakeni yamuva, i-phosphorylation yesayithi ethile ye-lysine ifinyelelwe kusetshenziswa ukusabela kwe-Staudinger-phosphite okukhethiwe ngamakhemikhali (Umfanekiso 3).
4. Myristoylation kanye palmitoylation
I-acylation ye-N-terminal enama-fatty acids ivumela ama-peptide noma amaprotheni ukuthi abophe ulwelwesi lwamaseli.Ukulandelana kwe-myridamoylated ku-N-terminal kwenza i-Src family protein kinases kanye ne-reverse transcriptase Gaq amaprotheni akhonjwe ukuze abophe ulwelwesi lwamaseli.I-Myristic acid yayixhunywe ku-N-terminal ye-resin-polypeptide kusetshenziswa ukusabela okujwayelekile kokuhlangana, futhi umphumela we-lipopeptide ungahlukaniswa ngaphansi kwezimo ezijwayelekile futhi uhlanzwe yi-RP-HPLC.
5. Glycosylation
Ama-Glycopeptides afana ne-vancomycin ne-teicolanin ama-antibiotic abalulekile ekwelapheni izifo ezibangelwa amagciwane amelana nemithi, kanti amanye ama-glycopeptides avame ukusetshenziselwa ukuvuselela amasosha omzimba.Ngaphezu kwalokho, njengoba ama-antigen amaningi amancane ane-glycosylated, kubaluleke kakhulu ukutadisha ama-glycopeptides ukuze kuthuthukiswe umphumela wokwelapha wokutheleleka.Ngakolunye uhlangothi, kutholakale ukuthi amaprotheni kulwelwesi lweseli lamaseli e-tumor akhombisa i-glycosylation engavamile, okwenza ama-glycopeptides adlale indima ebalulekile ocwaningweni lokuvikela umdlavuza nolwelwesi lwesimila.Ama-Glycopeptides alungiswa ngendlela ye-Fmoc/t-Bu.Izinsalela ze-glycosylated, njenge-threonine ne-serine, zivame ukwethulwa kuma-polypeptides nge-pentafluorophenol ester ecushiwe ama-fMOCs ukuvikela ama-amino acid e-glycosylated.
6. I-isoprene
I-Isopentadienylation yenzeka ezinsalela ze-cysteine ku-chain chain eduze kwe-C-terminal.Iphrotheni isoprene ingathuthukisa ukuhlangana kwe-cell membrane futhi yakhe ukusebenzisana kwamaprotheni-namaprotheni.Amaprotheni ane-Isopentadieated ahlanganisa i-tyrosine phosphatase, i-GTase encane, ama-molecule e-cochaperone, i-nuclear lamina, namaprotheni abopha ama-centromeric.I-Isoprene polypeptides ingalungiswa kusetshenziswa i-isoprene kuma-resin noma ngokwethula okuphuma ku-cysteine.
7. Ukuguqulwa kwe-polyethylene glycol (PEG).
Ukuguqulwa kwe-PEG kungasetshenziswa ukuthuthukisa ukuzinza kwe-protein hydrolytic, biodistribution kanye ne-peptide solubility.Ukwethulwa kwamaketanga e-PEG kuma-peptide kungathuthukisa izakhiwo zawo zemithi futhi kuvimbele i-hydrolysis yama-peptide ngama-proteolytic enzymes.Ama-peptide e-PEG adlula engxenyeni ye-glomerular capillary cross kalula kakhulu kunama-peptide avamile, kunciphisa kakhulu ukuphuma kwezinso.Ngenxa yempilo enwetshiwe esebenzayo yama-peptide e-PEG ku-vivo, izinga lokwelapha elijwayelekile lingagcinwa ngemithamo ephansi kanye nemithi ye-peptide engavamile.Nokho, ukuguqulwa kwe-PEG nakho kunemiphumela emibi.Inani elikhulu le-PEG livimbela i-enzyme ekonakaliseni i-peptide futhi linciphisa ukubophezela kwe-peptide kumamukeli okuqondiwe.Kodwa ukuhlobana okuphansi kwama-PEG peptides kuvame ukuxazululwa yi-half-life yawo ende ye-pharmacokinetic, futhi ngokuba khona emzimbeni isikhathi eside, ama-PEG peptides anethuba elikhulu lokumuncwa ezicutshini eziqondiwe.Ngakho-ke, ukucaciswa kwe-polymer ye-PEG kufanele kuthuthukiswe ukuze kube nemiphumela emihle.Ngakolunye uhlangothi, ama-peptide e-PEG anqwabelana esibindini ngenxa yokuncipha kokucaciswa kwezinso, okuholela ku-macromolecular syndrome.Ngakho-ke, ukuguqulwa kwe-PEG kudinga ukuklanywa ngokucophelela okukhulu lapho ama-peptide esetshenziselwa ukuhlolwa kwezidakamizwa.
Amaqembu avamile okulungisa ama-PEG modifiers angafinyezwa kanjena: Amino (-amine) -NH2, aminomethyl-Ch2-NH2, hydroxy-OH, carboxy-Cooh, sulfhydryl (-Thiol) -SH, Maleimide -MAL, succinimide carbonate - SC, succinimide acetate -SCM, succinimide propionate -SPA, n-hydroxysuccinimide -NHS, Acrylate-ch2ch2cooh, aldehyde -CHO (njenge-propional-ald, butyrALD), isisekelo se-acrylic (-acrylate-acrl), azido-azide, biotinyl - I-Biotin, i-Fluorescein, i-glutaryl -GA, i-Acrylate Hydrazide, i-alkyne-alkyne, i-p-toluenesulfonate -OTs, i-succinimide succinate -SS, njll. Okuphuma ku-PEG okune-carboxylic acid kungahlanganiswa ne-n-terminal amine noma amaketanga aseceleni e-lysine.I-PEG ecushwe yi-amino ingahlanganiswa ne-aspartic acid noma amaketango aseceleni e-glutamic acid.I-PEG engasebenzi kahle ingahlanganiswa ne-mercaptan yamaketanga aseceleni e-cysteine avikelwe ngokuphelele [11].Izilungisi ze-PEG zivame ukufakwa kusigaba esilandelayo (qaphela: i-mPEG ithi methoxy-PEG, CH3O-(CH2CH2O)n-CH2CH2-OH) :
(1) isilungisi se-PEG seketango eliqondile
mPEG-SC, mPEG-SCM, mPEG-SPA, mPEG-OTs, mPEG-SH, mPEG-ALD, mPEG-butyrALD, mPEG-SS
(2) isilungisi se-PEG esisebenza kabili
HCOO-PEG-COOH, NH2-PEG-NH2, OH-PEG-COOH, OH-PEG-NH2, HCl·NH2-PEG-COOH, MAL-PEG-NHS
(3) ukuhlanganisa i-PEG modifier
(mPEG)2-NHS, (mPEG)2-ALD, (mPEG)2-NH2, (mPEG)2-MAL
8. I-Biotinization
I-Biotin ingaboshwa ngokuqinile ne-avidin noma i-streptavidin, futhi amandla okubopha asondele ngisho nesibopho esivumelanayo.Ama-peptide anelebuli ye-Biotin avame ukusetshenziswa ku-immunoassay, i-histocytochemistry, ne-fluorescence-based flow cytometry.Amasosha omzimba anelebula e-antibiotin angasetshenziswa futhi ukubopha ama-peptide e-biotinylated.Amalebula e-Biotin avame ukunamathiselwa kuchungechunge oluseceleni lwe-lysine noma kutheminali engu-N.I-6-aminocaproic acid ivame ukusetshenziswa njengesibopho phakathi kwama-peptides ne-biotin.Ibhondi iyaguquguquka ekubopheni ku-substrate futhi ibopha kangcono ebukhoneni besithiyo se-steric.
9. Ukufakwa ilebula kwe-Fluorescent
Ukufakwa ilebula kwe-fluorescent kungasetshenziswa ukulandelela ama-polypeptide kumaseli aphilayo kanye nokutadisha ama-enzyme nezindlela zokusebenza.I-Tryptophan (Trp) iyi-fluorescent, ngakho-ke ingasetshenziselwa ukulebula kwangaphakathi.I-spectrum yokukhishwa kwe-tryptophan incike endaweni ezungezile futhi yehla ngokuncipha kwe-solvent polarity, impahla ewusizo ekutholeni ukwakheka kwe-peptide nokubophezela kwesamukeli.I-Tryptophan fluorescence ingacinywa nge-protonated aspartic acid ne-glutamic acid, enganciphisa ukusetshenziswa kwayo.Iqembu le-Dansyl chloride (Dansyl) linokukhanya okuphezulu kakhulu uma liboshwe eqenjini le-amino futhi livame ukusetshenziswa njengelebula le-fluorescent lama-amino acid noma amaprotheni.
Ukuguqulwa kwe-Fluorescence resonance Energy (FRET) kuyasiza ezifundweni zama-enzyme.Uma i-FRET isetshenziswa, i-polypeptide engaphansi ngokuvamile iqukethe iqembu elilebula le-fluorescence kanye neqembu le-fluorescence-quenching.Amaqembu e-fluorescent anelebula acinywa isicishi ngokudluliswa kwamandla okungewona we-photon.Lapho i-peptide ihlukaniswa ne-enzyme okukhulunywa ngayo, iqembu lamalebula likhipha i-fluorescence.
10. Ama-polypeptides ekheji
Ama-cage peptide anamaqembu okuvikela akhiphekayo avikela i-peptide ekubopheni i-receptor.Lapho ivezwe emisebeni ye-UV, i-peptide iyasebenza, ibuyisela ukuhlobana kwayo nesamukeli.Ngenxa yokuthi lokhu kuvula amehlo kungalawulwa ngokuya kwesikhathi, i-amplitude, noma indawo, ama-cage peptide angasetshenziswa ukutadisha ukusabela okwenzeka kumaseli.Amaqembu okuvikela asetshenziswa kakhulu ama-polypeptide ekheji amaqembu angu-2-nitrobenzyl kanye nokuphuma kwawo, angethulwa ekuhlanganiseni kwe-peptide ngokuphuma kokunye okuphuma ku-amino acid okuvikelayo.Ukuphuma kwe-amino acid okuye kwathuthukiswa i-lysine, i-cysteine, i-serine, ne-tyrosine.I-aspartate ne-glutamate derivatives, nokho, ayivamisile ukusetshenziswa ngenxa yokuthambekela kokuhamba ngebhayisikili ngesikhathi se-peptide synthesis and dissociation.
11. I-Polyantigenic peptide (MAP)
Ama-peptide amafushane ngokuvamile awavikeleki futhi kufanele ahlanganiswe namaprotheni okuthwala ukukhiqiza amasosha omzimba.I-Polyantigenic peptide (MAP) yakhiwe ngama-peptide amaningi afanayo axhunywe ku-lysine nuclei, engaveza ngokuqondile amandla aphezulu avikela umzimba futhi angasetshenziswa ukulungisa ama-couplets amaprotheni aphethe i-peptide.I-MAP polypeptides ingahlanganiswa ngokuhlanganisa kwesigaba esiqinile ku-MAP resin.Kodwa-ke, ukuhlangana okungaphelele kubangela amaketanga e-peptide alahlekile noma ancishisiwe kwamanye amagatsha futhi ngalokho akubonisi izici ze-MAP polypeptide yoqobo.Njengenye indlela, ama-peptide angalungiswa futhi ahlanzwe ngokuhlukana bese ehlanganiswa ne-MAP.Ukulandelana kwe-peptide okunamathiselwe kumongo we-peptide kuchazwe kahle futhi kubonakala kalula nge-mass spectrometry.
Isiphetho
Ukuguqulwa kwe-peptide kuyindlela ebalulekile yokuklama ama-peptide.Ama-peptide ashintshiwe ngamakhemikhali awakwazi nje ukugcina umsebenzi ophezulu wezinto eziphilayo, kodwa futhi agweme ngokuphumelelayo izithiyo ze-immunogenicity kanye nobuthi.Ngasikhathi sinye, ukuguqulwa kwamakhemikhali kunganikeza ama-peptide ngezakhiwo ezintsha ezinhle kakhulu.Eminyakeni yamuva nje, indlela yokusebenza kwe-CH ye-post-modification ye-polypeptides iye yathuthukiswa ngokushesha, futhi imiphumela eminingi ebalulekile ifinyelelwe.
Isikhathi sokuthumela: Mar-20-2023