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E ngoliloe ke Peptides Supplier Litšoantšo tsa Peptides Potso "ho tse latelang ke efe e sebelisoang ho theha li-peptide" hangata e hlaha ho e 'ngoe ea litlhophiso tse peli: ntho e kgethiloeng hangata moo condensation, hydrolysis, phapanyetso le esterification di qothisana lehlokoa bakeng sa karabo, kapa bothata ba benche moo equation ea buka e lokelang ho kopanngoa le protocol ea coupling e sebetsang lihora tse peli ho DMF. Litlhophiso ka bobeli li putsa phapang e tšoanang. Karabelo e bakang tlamo ea peptide ke ho theha bond ea amide ka nucleophilic acyl substitution at activated carboxyl carbon.. The condensation equation lithuto tse ngata li hatisoa - carboxyl mmoho le amine e fanang ka amide le metsi - ke net stoichiometry ea phetoho eo., ha se mokhoa oo motho a o tsamaisang joalo ka ha ho ngoliloe.
Ho latela maikutlo a mahlano a fosahetseng ka lebaka la ho putlama ha phapang eo, 'me e' ngoe le e 'ngoe e lokisoa ka tlase ka mochine oa eona, bopaki le maemo a moeli. Karolo ea botšelela e nka mohlala o fapaneng oo potso e sa sebetseng ho hang: li-non-ribosomal peptide synthetases, moo ho se nang reagent e kopanyang 'me ho se na potoloho ea sehlopha se sireletsang. Ntle le ha ho boletsoe ka hosele, tleleime e 'ngoe le e 'ngoe mona e itšetlehile ka lik'hemik'hale (e seng ribosomal) tlhahiso ea li-peptide, mme mehlala ya dipalo e ntshitswe ho tatellano le maemo a itseng a boletsweng, eseng ho tsoa ho peptide synthesis ka kakaretso.
Maikutlo a fosahetseng 1: "Condensation equation ke karabelo eo u e tsamaisang"
Phoso ke ho bala equation e leka-lekaneng joalo ka risepe. Ho kopanya asiti ea mahala ea carboxylic le amine ea mahala ho fana ka letsoai la ammonium carboxylate, 'me tekatekano bakeng sa ho etsoa ha amide ka ho toba metsing e itšetlehile hantle ka letsoai ho e-na le amide. Ho futhumatsa ho khanna metsi ha se tharollo e sebetsang ha balekane ba nkile liketane tse ka mahlakoreng tse arabelang, epimerise kapa ho bola tlasa maemo ao.
Se hlileng se etsahalang ka ho kopanya ke tatellano ea likarolo tse peli. Ntlha ea pele, karolo ea carboxyl e fetoloa ho ba mefuta e mengata ea electrophilic - O-acylisourea e tsoang ho carbodiimide e kang DCC., DIC kapa EDC, ester e sebetsang ho tsoa HOBt, HOAT kapa Oxyma, kapa mofuta o hlahisitsoeng ke uronium/phosphonium. Ebe litlhaselo tsa amine tse kentseng carbonyl, mme sehlopha sa tlatsetso kapa se tlohang se a tloha. OpenStax Organic Chemistry §26.7, e hatisitsoeng ka September 20, 2023, e hlalosa hantle sena: amide e bonolo e ka etsoa ka ho phekola amine le asiti ea carboxylic ka carbodiimide, empa peptide synthesis e thata ho feta hobane li-amide tse ngata tse fapaneng li tlameha ho theha ka tatellano e itseng ho e-na le ho iketsahalla feela. Tataiso ea ho kopanya-reagent ea Bachem e thathamisa malapa a activation a sebelisoang - acid halides, azides, li-anhydride tsa symmetrical le tse tsoakiloeng, carbodiimides e nang le li-additives kapa ntle le eona - e leng khopotso ea bohlokoa ea hore "ts'ebetso" ke sehlopha sa ts'ebetso., eseng reagent e le 'ngoe.
Meeli e bohlokoa bakeng sa likarabo tsa litlhahlobo le bakeng sa moralo oa benche. Haeba potso e botsa hore na ke sehlopha sefe sa karabelo se etsang bond ea peptide, karabo ke ho thehoa ha amide ka nucleophilic acyl substitution at activated carboxyl carbon.; condensation e hlalosa tekanyo ea athomo ka kakaretso. Haeba potso e botsa hore na ho kenya eng ka botlolong, ha e le equation kapa lebitso la sehlopha ha le lekane - reagent, tlatsetso, solvent, motheo le mocheso kaofela li tlameha ho boleloa, 'me likhetho tseo ke tseo likarolo tse setseng li buang ka tsona.
Maikutlo a fosahetseng 2: "Lihlopha tsa ho sireletsa ke boikhethelo haeba u hlokolosi"
Phoso ke ho nka ts'ireletso joalo ka ntlafatso ea lihlahisoa. Ke tlhokahalo ea khetho. Amino acid e 'ngoe le e' ngoe e hlahisa bonyane α-amine le α-carboxyl, 'me boholo ba bona ba na le nucleophile ea lehlakoreng le leng - the thiol of cysteine, imidazole ea histidine, ε-amine ea lysine, hydroxyl ea serine, threonine kapa tyrosine. Ntle le tšireletso ea orthogonal, motsoako o kopanyang o na le li-nucleophile tse ngata tse qothisanang lehlokoa le carboxyl e tšoanang, mme kabo ya sehlahiswa ke ya dipalopalo ho fapana le ho hlaloswa ka tatelano.
Mohlala o sebetsang oa OpenStax oa Ala-Leu o etsa konkreite ea logic: ho kopanya alanine le leucine, amine ea alanine le carboxyl ea leucine li sirelelitsoe, amide e lakatsehang e thehoa ka EDC kapa DCC, ebe lihlopha tse sireletsang lia tlosoa - mehato e mehlano bakeng sa dipeptide. Mohloli o tšoanang o hlokomela hore lihlopha tsa amino hangata li nkoa e le Boc (e tlosoa ke acid e matla e khuts'oane joalo ka TFA) kapa, hangata haholo, joalo ka Fmoc (tlosoa ke 20% piperidine ka DMF), athe lihlopha tsa carboxyl hangata li tšoaroa joalo ka methyl kapa benzyl esters. The Wikipedia peptide synthesis kakaretso liforeimi ntlha e ts'oanang boemong ba leano: merero e sireletsang-sehlopha hangata e ea hlokahala ho thibela maikutlo a mahlakore ka liketane tse mahlakoreng, 'me motsoako oa lik'hemik'hale ka tloaelo o qala ho C-terminus ebe o fetela ho N-terminus.
Boemo ba moeli ke 'nete empa bo patisane. Chemoselective ligation e atamela le e khutšoanyane, likhechana tse sebetsang ka mokhoa o ikhethileng li ka fokotsa moroalo oa tšireletso, hobane balekane ba arabelang ba khethoa e le hore ho ka kopana para e le 'ngoe feela. Ha ba e tlose bakeng sa kopano ea ketane ea stepwise, moo ketane e ntseng e hola le mesaletsa e kenang ka bobeli e tsamaisang litsi tse ngata tse sebetsang. Tlhahlobo e sebetsang: haeba chromatogram e tala e bonts'a lere ea litlhōrō tse arohaneng ke masala a le mong ho e-na le sehlahisoa se le seng se ka sehloohong., bothata hangata ke ho khetha, ha reagent e sa lekaneng.
Maikutlo a fosahetseng 3: "Activation le coupling ha e nke lehlakore"
Phoso ke ho nka hore coupling reagent e ama feela hore na lihlahisoa tsa mofuta ofe. E boetse e ama mefuta ea stereoisomer. The Limolek'hule tlhahlobo ea Duengo et al. (2023; 28:8017) e hlalosa litsela tse peli tsa motheo tsa ho lahleheloa ke tlhophiso: ho ntšoa ka ho otloloha ha α-proton ho fana ka enolate e tsoang sefahlehong ka seng, le ho thehoa ha oxazol-5(4H)-ngoe ho tsoa masalla a kentsweng tshebetsong, e leng tsela e atileng haholo ha carboxyl e kentswe tshebetsong. Litsela ka bobeli li kopana ka temoso e le 'ngoe - masala a kentsoeng a carboxyl ke sebaka se kotsi ka ho fetisisa sa potoloho., mme kotsi e behilwe ke matla a motheo, steric bongata le hore na mefuta e kentsoeng e lula nako e kae pele amine e fihla.
Kakaretso ea tlhahlobo ea data ea papiso e loketse ho qotsoa ka boholo ba eona bo sa fetoheng. Ka mokhoa o tiileng oa motsoako oa Gly-Phe-Pro-NH₂, tlaleho ea epimerization e ne e 29.8% ka EDC/HOAT, 24.1% le EDC·HCl/HOAt le 4.2% ka DIC/HOAT. Tsena ke lipalo tsa tatellano ea mohlala o le mong tse hlophisitsoeng ho tsoa lithutong tsa mantlha tse boletsoeng; they do not establish a general ranking of EDC against DIC across sequences, and the review elsewhere notes that in some comparisons the difference between EDC and DCC was marginal. Motsotsoana, separately attributed data point comes from vendor technical documentation comparing histidine derivatives: Fmoc-His(Trt)-OH coupled with HCTU/6-Cl-HOBt/DIPEA gave 1% racemisation without preactivation and 7.8% with five minutes of preactivation, while Fmoc-His(MBom)-OH under the same preactivation gave 0.3%. That is a vendor comparison of specific derivatives and conditions, not a claim that Trt protection is unsuitable for histidine.
The corrected statement is conditional: reagent, tlatsetso, base and preactivation time each shift the epimerisation outcome, and the size of the shift depends on the residue being activated and on how slow the coupling is. Likopano tse liehang tlas'a maemo a mantlha ke tsona moo mefuta e kentsoeng e nang le nako ea ho palama baesekele. Ka hona botšepehi ba stereochemical bo tlameha ho lekanyetsoa - ka tlhahlobo ea chiral, Reagente ea Marfey, kapa chromatography ea boemo bo holimo e khona ho rarolla diastereomer - ho fapana le ho tsoa ho khokahano e shebahalang e hloekile.
Maikutlo a fosahetseng 4: "Nomoro e le 'ngoe ea bohloeki e u bolella hore tatellano e nepahetse"
Phoso ke ho bala peresente ea bohloeki joalo ka polelo ea boitsebiso. HPLC area% e araba potso e moqotetsane: ea lintho tse ileng tsa lokoloha 'me e ile ea fumanoa ka wavelength e khethiloeng, karolo efe e neng e le tlhorong e kgolo? Ha e bolele hore na tlhoro eo ke efe. Letoto la ho hlakolwa, diastereomer ho tsoa ho epimerization, kapa sehlahisoa se entsoeng ka aspartimide se ka kopana le sepheo tlas'a sete e le 'ngoe ea maemo a gradient., le ea Limolek'hule hlahloba lintlha ka ho hlaka hore lihlahisoa tsa epimerised li na le litšobotsi tse tšoanang tsa 'mele, ke ka baka leo ho leng thata ho hlwekisa. The ChemPep Fmoc SPPS tataiso ea tekheniki e etsa ntlha e bapileng ho tloha lehlakoreng la phallo ea mosebetsi: Maemo a "tloaelehileng" a ho kopanya a ka fana ka kopanyo e sa phethahalang le lihlahisoa tse ling tse kenyelletsang li-epimers., li-peptide tse fokolang le lihlahisoa tsa Fmoc cleavage pele ho nako.
Mass spectrometry e koala karolo ea lekhalo empa eseng kaofela ha eona. Boima bo hlokometsoeng bo tsamaellanang le boima bo baliloeng bo netefatsa sebopeho, eseng boemo ba tatellano - tatellano e qhetsoeng e nang le mesaletsa e tšoanang e fana ka boima bo tšoanang. Boitsebiso ba boemo bo hloka tandem MS, Ho senyeha ha Edman, kapa NMR bakeng sa li-peptide tse khutšoane. Ka thoko, foromo ea counterion (TFA ka ho sa feleng ho li-catalogs tse ngata, ka acetate, chloride kapa ammonium phapanyetsano e fumaneha), metsi le net peptide content ke bongata bo fapaneng le bohloeki, le a 98% Palo ea HPLC ha e bue letho ka e 'ngoe ea tsona.
Moeli ke hore litekanyetso tsa ho amoheloa li itšetlehile ka tšebeliso e reretsoeng. Sesebelisoa sa boemo ba lipatlisiso bakeng sa tlhahlobo ea invitro le thepa e reretsoeng mosebetsi oa pele ho kliniki ha e na litlhaloso tse tšoanang., mme setifikeiti sa sehlopha sa lipatlisiso ha se tlhaloso ea meriana. Tafole e ka tlase ke moralo oa tlhahlobo bakeng sa ho etsa qeto ea hore na ke lets'oao lefe la tlhahlobo le tla senola ho hloleha hofe; ha se mokhoa o tiisitsoeng, le meeli ea ho lemoha e itšetlehile ka kholomo, gradient, bolelele ba maqhubu le sesebedisoa se sebedisoang.
Tafole: Sepheo sa ho arabela, litloaelo tsa lehlakore tse ikhethang, le bopaki ba tlhahlobo
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Mohato oa ho itšoara |
Morero |
Phetoho e ka bang teng |
Bopaki ba ho hlahloba |
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Ts'ebetso ea carboxyl |
Fetola carboxyl hore e be mofuta oo amine e ka o hlaselang |
Epimerization ka oxazol-5(4H)-ngoe; Ho thehoa ha N-acyl urea ka carbodiimides; karabelo ya pele ho nako ka di-nucleophiles tsa mahlakoreng |
Diastereomer tlhoro ho chiral kapa high-resolution HPLC; boima bo eketsehileng bo tsamaellanang le acyl urea ho LC-MS |
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Amide bond sebopeho (Kopanya) |
Eketsa ketane ka masala a le mong |
Khokahano e sa fellang e fanang ka tatelano ea ho hlakolwa; tlatsetso habeli; guanylation by-products tse nang le li-reagents tsa uronium |
−AA ho fetoha ha boima ho LC-MS; +AA e fetoha ka bongata; boima ba guanidinium |
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Fmoc dep Fektheri e bitsoang Peptides tšitiso |
Hlahisa Nα-amine bakeng sa potoloho e latelang |
Tšireletso e sa phethahalang; sebopeho sa aspartimide ho Asp-Xxx tlasa maemo a mantlha; sebopeho sa diketopiperazine kamora ho sireletsoa ha li-intermediate tse itseng tsa dipeptidyl-resin |
Residual Fmoc ka UV kapa MS; +18/−18 Lipaterone tsa Da ho tloha ho lesale la aspartimide ho bula ho α- le lihlahisoa tsa β-aspartyl; tahlehelo ea boima ba dipeptide |
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Tšireletso ea mahlakoreng Peptide Reconstitution le cleavage |
Lokolla peptide 'me u tlose lihlopha tse sireletsang mahlakoreng |
Sekhahla se sa fellang se siea masala a sirelelitsoeng; alkylation ea Cys, Met or Trp by cleavage-cocktail cations |
Liphetoho tse ngata tse tsamaellanang le lihlopha tse bolokiloeng tse sireletsang; eketsa bongata |
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Ho itšehla thajana ho qetela |
Tlosa letsoai, counterion exchange, lyophilise |
TFA e setseng; metsi a mangata; litaba tse tlase tsa peptide |
Ion chromatography bakeng sa TFA e setseng; Karl Fischer bakeng sa metsi; tlhahlobo ea mantlha kapa ea bongata ba amino acid bakeng sa peptide ea net |
Tlhaloso: Moralo oa tlhahlobo, ha se protocol e netefalitsoeng. Letšoao le leng le le leng le bontša seo u lokelang ho se batla; hore na mokhoa o fanoeng o ka o lemoha boemong bo loketseng ho itšetlehile ka meeli ea mokhoa oo.
Maikutlo a fosahetseng 5: “Haeba kopano e sebetsa, karolo e setseng e hantle”
Phoso e akaretsa ho tloha mohatong o le mong o atlehileng ho ea ho sehlopha sohle. Katleho ea ho kopanya e itšetlehile ka tatellano, le mekhoa ea ho hlōleha eo e leng ea bohlokoa ka ho fetisisa e bonahala e le masala a sieo ho e-na le ho ba le lihlahisoa tse tlaase. The Limolek'hule tekolo botjha e bolela paterone ka kotloloho: khokahanyo le ts'ireletso ea ts'ireletso e fokotseha ka bolelele ba ketane ea peptide, ho hlahisa li-amino acid le tatellano e sa phethahalang, le likhechana tse kentsoeng C-terminal tse nang le karolo ea mahala ea N-terminal li eketsa haholo monyetla oa ho phatloha. Tataiso ea ChemPep e eketsa corollary e sebetsang ea hore peptide aggregation nakong ea synthesis ke sesosa se seholo sa mathata., ho thata ho bolela esale pele, e itshetlehile ka tatelano, mme ha e na tharollo ea bokahohle - ke ka lebaka leo protocol e tšoanang e ka sebetsang ka mokhoa o hloekileng ka tatellano e le 'ngoe' me ea hlōleha ho e 'ngoe ea bolelele bo tšoanang..
Likarolo tse tharo tsa tatellano li hloka tlhokomelo e khethehileng. Metsotsoana e telele ea hydrophobic e khothalletsa ho kopana ha resin le ho kopanya ho sa fellang, e hlahang joalo ka tatellano ea ho hlakolwa. Asp-Xxx motifs e tloaetse ho etsoa ha aspartimide tlasa maemo a mantlha a ts'ireletso ea Fmoc., ho fana α- le lihlahisoa tsa β-aspartyl ka mor'a ho bula lesale. Li-intermediate tse ling tsa dipeptidyl-resin li ka tsamaela ho diketopiperazine kamora ho sireletsoa., ho fokotsa ketane pele ho kopanyo e latelang. Ha ho le e 'ngoe ea tsena e ipolelang e le chai e tlase ea ho kopanya ka nako eo; e 'ngoe le e 'ngoe e hlaha hamorao e le litšila tse fetohang ka bongata.
Taba e lokisitsoeng ke hore boleng ba batch bo thehiloe ke data e khethehileng ea tlhahlobo ho pholletsa le mekhoa ea orthogonal., 'me bopaki bo loketseng bo itšetlehile ka bolelele ba tatellano, hydrophobicity le dikahare tsa phetoho. E khuts'oane, hydrophilic, peptide e sa fetoloang le peptide ea 40-residue hydrophobic e nang le li-disulfide tse peli ha e fane ka tumello ea tlhahlobo e tšoanang ea tokollo., 'me ho li nka li lekana ke kamoo tlhaloso e shebahalang e le thata e qetellang e sa bue hakaalo.
Mohlala oa counter: moo potso e sa sebetseng teng
Non-ribosomal peptide synthetases ke moo "karabelo efe e hlahisang peptide" e nang le karabo e fapaneng haholo., le moo khemikhale-synthesis foreimi e sa feteleng. The tlhahlobo ea sebopeho sa enzymology ea NRPS e hlalosa li-enzyme tsa modular moo mojule ka mong o kenyelletsang masala a le mong. Amino acid e sebelisoa e le acyl-adenylate ke sebaka sa adenylation, ebe e fetisetsoa e le thioester ho letsoho la phosphopantetheine la protheine e tsamaisang peptidyl, 'me bond ea peptide e thehoa sebakeng sa condensation se nang le sepheo se bolokiloeng sa HHxxxDG.. Sebaka sa terminal sa thioesterase se lokolla sehlahisoa.
Liphapang tse tharo li etsa hore moralo oa lik'hemik'hale o se ke oa sebetsa. Ha ho na exogenous coupling reagent - ts'ebetso ke enzymatic mme e itšetlehile ka ATP. Ha ho na potoloho ea Nα e sireletsang, hobane protheine e tsamaisang thepa e tšoara substrate ebe e e isa sebakeng se nepahetseng. Mme stereochemistry e behiloe ke khetho ea enzyme ho fapana le maemo a ts'ebetso, ke kamoo litsela tsa NRPS li kenyelletsang li-amino acid tsa D-amino ntle le bothata ba epimerization Limolek'hule tlhahlobo. E ngotsoe Peptides Wholesale
Tlhahlobo e hlakile mabapi le meeli ea eona, 'me meeli eo e lokela ho tsamaea le mohlala oa counter. Mehaho e fumanehang ea condensation-domain ha e na li-ligand tsa tlhaho sebakeng se sebetsang, kahoo mesebetsi e nepahetseng ea masala a sebaka se sebetsang ho tlamang le ho kenya letsoho ho sa ntse ho tla thehoa, 'me matla a libaka tsa condensation ha a rarolloe. Ka hona mohlala oa counter o tšehetsa phapang ea sebopeho - kopano e entsoeng ka enzymatic khahlano le khokahanyo ea lik'hemik'hale e tsamaisoang ke reagent - eseng ho lekana ka botlalo pakeng tsa tse peli..
Seo u ka se botsang ha u sa khone ho etsa tlhahlobo ka bouena
Bakeng sa babali ba laelang synthesis ho e-na le ho e tsamaisa, maikutlo a fosahetseng a mahlano a fetoha lethathamo le lekhutšoanyane la lintho tseo u ka li botsang. Taba ha se ho batla ntho e 'ngoe le e' ngoe bakeng sa peptide e 'ngoe le e' ngoe, empa ho bapisa bopaki le mekhoa ea ho hloleha tatellano e hlile e pepesitsoe.
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HPLC e khethehileng haholo e nang le mekhoa ea mekhoa. Kholomo, gradient, wavelength le nako ea ho matha, kahoo palo ea bohloeki e ka hlalosoa ho e-na le ho nkoa ka tšepo. Mokhoa oa ho kopanya li-peptide
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MS e nang le boima bo hlokometsoeng le bo baloang. Bobeli litekanyetso, kahoo papiso e ka hlahlojoa.
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Sebaka sa phetoho le k'hemistri. E leng masalela a nang le leibole, sehlopha sa tshireletso se sebedisoang, le hore na phetoho ea bongata e lebelletsoeng ke efe.
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Foromo ea ho hanyetsa. TFA ka ho sa feleng ho li-catalogs tse ngata, ka acetate, chloride kapa ammonium phapanyetsano e fumaneha ka kopo.
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Teko ea stereochemical kapa litaba moo tatellano e fanang ka eona. Tlhahlobo ea Chiral bakeng sa masala a tloaelehileng a ho epimerization; litaba tsa peptide ka tlhahlobo ea amino acid ea mantlha kapa ea bongata moo peptide e bohlokoa ho feta sebaka%.
MOL Changes e sebelisana le HPLC le MS data ea tokollo mme e fana ka boikhethelo ba litaba, tlhahlobo ea endotoxin le tlhahlobo ea sterility bakeng sa tatellano moo lintlha tseo li sebetsang teng; tatelano e hlalositsoeng le ts'ebeliso e reriloeng li lekane ho kopa qotso e tsamaellanang le lintlha. Lisebelisoa tsa mofuta ona tsa lipatlisiso li etselitsoe mosebetsi oa laboratori mme ha se sehlahisoa sa meriana kapa thepa e lokiselitsoeng ho sebelisoa ke batho..
Tsebiso: MOL Changes ke moetsi oa li-peptide tsa khoebo mme o na le thahasello ea khoebo holima maemo a boleng a boletsoeng mona.
Mehloli
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OpenStax, Organic Chemistry, §26.7 "Peptide Synthesis,” e hatisitsoeng ka September 20, 2023. https://openstax.org/books/organic-chemistry/pages/26-7-peptide-synthesis
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Duengo, S., Muhajir, M. I., Al-Anshori, J., le al. "Racemization / Epimerization ho Peptide Synthesis." Limolek'hule 2023, 28:8017. https://pmc.ncbi.nlm.nih.gov/articles/PMC10745333/
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"Structural Biology of Non-Ribosomal Peptide Synthetases." PMC4760355. https://pmc.ncbi.nlm.nih.gov/articles/PMC4760355/
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ChemPep, "Fmoc Solid Phase Peptide Synthesis" (barekisi litokomane tsa tekheniki, e sebelisetsoa ho hlalosa mokhoa oa ho sebetsa feela). https://chempep.com/fmoc-solid-phase-peptide-synthesis/
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Bachem, "Peptide Synthesis e sebetsang hantle: Tataiso ea ho Coupling Reagents le Additives” (barekisi litokomane tsa tekheniki, e sebelisetsoa tlhaloso ea sehlopha sa reagent feela). https://www.bachem.com/knowledge-center/efficient-peptide-synthesis-a-guide-to-coupling-reagents-additives/
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Histidine racemisation papiso (1% / 7.8% / 0.3%): papiso ea tekheniki ea morekisi ea Fmoc-Derivatives ea hae tlasa HCTU/6-Cl-HOBt/DIPEA, e sebelisoa e le sebaka sa data se ikhethileng, eseng joalo ka maemo a akaretsang.
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Motsamaisi oa Lab, "Litšebelisano tsa DNA-Peptide li theha mekhoa e rarahaneng e ka 'nang eaba e thusitse ho fetola baeloji" - e hlahlobiloe ebile ha e sebelisoe, hobane ha e sebetsane le sebopeho sa bond ea lik'hemik'hale tsa peptide.
LBH
Ke sehlopha sefe sa karabelo se etsang tlamo ea peptide, le hore na ho kenya tshebetsong ho eketsa eng? Ho theha maqhama a Amide ka ho nkela sebaka sa nucleophilic acyl ho khabone ea carboxyl. Activation e fetola carboxyl hore e be electrophile e betere - e leng O-acylisourea, ester e sebetsang, kapa mofuta o tsoang ho uronium / phosphonium - kahoo amine e ka hlasela tlas'a maemo a bonolo. Condensation equation ke tekano ea athomo e felletseng ea phetoho eo, eseng mokhoa.
Na Fmoc kapa Boc e betere bakeng sa tatelano e tloahelehileng ea ho matha? E itšetlehile ka masala le maemo a ho kopanya, mme bopaki ha bo tshehetse karabo e akaretsang. The Limolek'hule tlhahlobo e hlokomela hore ts'ireletso ea Fmoc e sebelisa motheo, e amanang le masalla a botlaaseng, ha chemistry ea Boc e sebelisa asiti; empa tlhahlobo e tšoanang e bontša hore coupling reagent, khetho ea tlatsetso le ea motheo hangata e laola sephetho sa epimerization. Bakeng sa tatelano e itseng, karabo e sebetsang ke ho tseba masala a kotsi e kholo le ho kopa mokhoa oa ho hlahloba o tla lemoha li-epimers tsa bona..
Na ho ka ba le peptide 98% e hloekileng ka HPLC mme e ntse e le molek'hule e fosahetseng? Ee. Area% e hlalosa karoloana ea lintho tse bonoang tlhorong e kholo, eseng seo tlhoro eo e leng sona. Letoto la ho hlakolwa, diastereomer kapa co-eluting by-product e ka laola chromatogram tlasa maemo a le mong.. Boitsebiso bo hloka MS hammoho le mokhoa o fanang ka boitsebiso ba boemo, le dikahare, counterion le metsi ke litekanyo tse arohaneng hape.
Na karabelo ea condensation e boetse e hlalosa kamoo lisele le li-enzyme tsa NRPS li etsang li-peptide? Che. Ribosomal le non-ribosomal biosynthesis ka bobeli li theha maqhama a amide, empa ka mekhoa ea template ea enzymatic - ts'ebetso ea acyl-adenylate, thioester tethering ho protheine e tsamaisang thepa, le catalysis ea "condensation-domain" - ha ho na reagent ea ho kopanya e sa tloaelehang ebile ha ho na potoloho ea sehlopha se sireletsang. Tekanyo ea athomo ea nete e tšoana; k'hemistri e e fihlelang ha e joalo.
Phatlalatso ea mohatisi: MOL Changes e fana ka litšebeletso tsa peptide mme e na le thahasello ea khoebo sehloohong sena. Mekhahlelo ea litšebeletso le lipolelo tsa bokhoni ke likopo tsa bafani ba thepa; ho fumaneha le ts'ebetso bakeng sa moaho o itseng o hloka bopaki bo ikhethileng ba projeke.
