GLP-1(link:https://www.bloomtechz.com/synthetic-chemical/peptide/glp-1-peptide-cas-87805-34-3.html) yog ib qho tshuaj hormone polypeptide uas muaj 30 amino acids. Nrog rau qhov kev tshawb fawb tob ntawm GLP-1, ntau thiab ntau cov txheej txheem hluavtaws tau tsim. Kab lus no yuav qhia txog cov txheej txheem kev sib txuas uas paub tam sim no ntawm GLP-1.
Txoj kev 1, khoom theem synthesis:
Solid-theem synthesis yog ib txoj hauv kev siv dav rau peptide thiab protein synthesis, thiab kuj tseem siv rau kev sib txuas ntawm GLP-1. Nyob rau hauv cov khoom-theem synthesis, cov qauv tseem ceeb yog tsim los ntawm kev sib txuas thawj cov amino acid rau cov resin. Tom ntej no, tom ntej no amino acid yog ntxiv nyob rau hauv ib theem zuj zus thiab chemically reacted nrog ib tug tsim nyog condensing neeg sawv cev. Thaum kawg, cov khoom lag luam tuaj yeem tau txais los ntawm cleaving polypeptide los ntawm cov resin.
Qhov tseem ceeb ntawm cov khoom siv-theem synthesis yog tias nws ua rau lub automation thiab loj-scale zus tau tej cov peptide synthesis. Cov txheej txheem tseem ceeb tam sim no-theem synthesis suav nrog Fmoc thiab Boc. Ntawm lawv, Fmoc txoj kev siv N-Fmoc tiv thaiv pab pawg los tiv thaiv peptide, thaum Boc txoj kev siv tert-butyloxycarbonyl los tiv thaiv pawg carboxyl.
Txoj kev 2, kua theem synthesis:
Liquid-theem synthesis yog ib txoj hauv kev ntawm peptide synthesis uas cov reactants muab tso rau hauv cov kua theem rau cov tshuaj tiv thaiv. Qhov kom zoo dua ntawm cov kua-theem synthesis yog tias cov tshuaj tiv thaiv mob me me thiab tsim nyog rau kev hloov kho cov qauv tshuaj rhiab heev. Txawm li cas los xij, vim muaj ntau cov reactants, cov txheej txheem purification yog qhov nyuaj. Cov tshuaj tiv thaiv hauv cov kua theem synthesis muaj xws li:
1. Cov tshuaj tiv thaiv condensation:
Cov tshuaj tiv thaiv condensation yog ib qho ntawm cov tshuaj tiv thaiv tseem ceeb tshaj plaws hauv peptide synthesis, uas yog, pawg carboxyl pib los ntawm cov neeg ua haujlwm condensing xws li DCC thiab HOBt txuas nrog cov amino pawg amino acid los ntawm acylation cov tshuaj tiv thaiv. Cov tshuaj tiv thaiv mob me me thiab cov txiaj ntsig siab.
2. Kev tshem tawm cov tshuaj tiv thaiv:
Kev tshem tawm cov tshuaj tiv thaiv yog txo cov methionine rau dithiol los ntawm NaBH4 thiab lwm cov tshuaj txo qis, ua rau nws tsis ua haujlwm. Cov tshuaj tiv thaiv yuav tsum tau ua raws li cov xwm txheej yooj yim.
3. Tshem tawm cov pab pawg tiv thaiv:
Vim muaj cov haujlwm sib txawv ntawm cov amino acids hauv cov saw peptide, cov pab pawg tiv thaiv sib txawv yuav raug siv los tiv thaiv. Tom qab cov synthesis tiav lawm, pawg tiv thaiv yuav tsum tau muab tshem tawm. Rau txoj kev Fmoc, piperidine feem ntau yog siv los tshem tawm Fmoc; Thaum rau txoj kev Boc, TFA yog siv los tshem tawm Boc.
Txoj kev 3, chemical synthesis:
GLP-1 yog ib qho tshuaj hormone polypeptide nrog cov dej num tseem ceeb hauv kev lom neeg. Nws cov synthesis tuaj yeem pom tau los ntawm ntau txoj hauv kev, ntawm cov tshuaj synthesis yog ib txoj hauv kev siv ntau tshaj plaws. Qhov kom zoo dua ntawm cov tshuaj synthesis yog tias nws tuaj yeem tsim cov khoom lag luam ntshiab heev, uas tsim nyog rau kev tsim khoom loj. Cov txheej txheem tshuaj synthesis thiab cov kauj ruam ntxaws ntawm GLP-1 yuav qhia hauv qab no.
1. Synthetic txoj kev thiab tiv thaiv pawg xaiv:
GLP-1 molecule muaj 36 amino acids, suav nrog 21 L-type thiab 15 D-type amino acids. Ua ntej nqa tawm cov synthesis, nws yog ib qho tsim nyog yuav tau xaiv ib txoj hauv kev uas tsim nyog thiab xaiv cov pab pawg tiv thaiv raws li cov khoom siv hluavtaws. Fmoc solid-phase synthesis feem ntau yog siv rau automated loj-scale synthesis. Txoj kev no siv N-9-fluoromido carboxyl kev tiv thaiv (N-Fmoc) ua pab pawg tiv thaiv, thiab tseem yuav tsum xaiv cov pab pawg tiv thaiv thib ob uas tsim nyog (xws li tert-butyl lossis methyl) txhawm rau tiv thaiv cov chaw tshwj xeeb. Txhua zaus ntxiv cov amino acid tshiab, Fmoc tiv thaiv pab pawg yuav tsum tau muab tshem tawm ua ntej, thiab tom qab ntawd ntxiv cov tshuaj tiv thaiv txuas ntxiv ntawm cov amino acid ntxiv.
2. Synthesis ntawm cov tub ntxhais amino acid ib ntus:
Cov txheej txheem tseem ceeb ntawm GLP-1 muaj 21 amino acids, suav nrog ib qho tseem ceeb serine thiab plaub prolyl-glutamic acid dipeptide sequences. Hauv cov txheej txheem synthesis, cov kev sib txuas ntawm cov tub ntxhais kawm tuaj yeem muab faib ua cov kauj ruam hauv qab no:
2.1. Ntxiv acetic acid carbamate (Fmoc-NH-CH2CO2Et) thiab 2-Cl-Trt-Cl rau cov khoom siv hluavtaws hluavtaws, thiab ua cov tshuaj tiv thaiv condensation nrog DIC/NMM coupling tus neeg sawv cev.
2.2. Tshem tawm Fmoc tiv thaiv pab pawg los ntawm kev tiv thaiv pab pawg cov tshuaj tiv thaiv.
2.3. Ntxiv cov amino acid tom ntej, rov ua cov kauj ruam 1 thiab qib 2 hauv ib ntus kom txog thaum cov tub ntxhais sib txuas ua ke.
2.4. Tsim cov qauv pentapeptide ntawm cov khoom theem resin. Ntxiv cov acetalization reagent rau cov khoom-theem resin, hnov mob nrog N-terminal recognition tus neeg saib xyuas (xws li HBTU), ntxiv sab saw tiv thaiv pawg ntawm serine ua tus pab txo tus neeg sawv cev, thiab tom qab ntawd tshem tawm Fmoc pawg tiv thaiv.
2.5. Nyob rau hauv lub catalysis ntawm Bacillus subtilis transferase (ProTide), cov qauv pentapeptide tau txais kev sib pauv tshuaj tiv thaiv nrog cov precursor ntawm serine iodoacetate.
3. Synthesis ntawm seem amino acid ib ntus:
Tom qab ua tiav cov synthesis ntawm cov tub ntxhais ua ntu zus, nws yog ib qho tsim nyog yuav tsum tau ntxiv cov amino acids ntxiv, suav nrog L- thiab D-hom amino acids. Qhov sib ntxiv ntawm cov amino acids no yuav tsum tau pib los ntawm cov txheej txheem tseem ceeb, ntxiv cov amino acid tom ntej hauv ib ntu, thiab siv tus neeg sawv cev condensing coj los ua cov tshuaj tiv thaiv kom txog thaum ua tiav GLP-1 polypeptide molecule yog synthesized. Thaum lub sijhawm txheej txheem no, nws tseem yuav tsum xaiv ib pab pawg tiv thaiv tsim nyog raws li qhov yuav tsum tau ua, thiab ua cov kauj ruam ntawm cov tshuaj tiv thaiv, tshem tawm cov pab pawg tiv thaiv thiab ntxiv cov amino acid nyob rau hauv ib ntus.
4. Sodium hydroxide kev kho mob:
Tom qab tag nrho cov amino acids tau ntxiv lawm, ib qho kev sib txuas tsis tiav peptide saw yog tsim los ntawm cov khoom siv-theem resin thiab yuav tsum tau ua tiav los ua kom tiav peptide molecule. Ua ntej, cov peptide unformed yuav tsum tau hydrolyzed los ntawm sodium hydroxide, thiaj li hais tias C-terminal carboxyl pab pawg neeg Ameslikas txuas rau cov resin yog detached los ntawm resin, thiab pab pawg neeg tiv thaiv yog detached nyob rau hauv dej. Tom qab hydrolysis cov tshuaj tiv thaiv, cov khoom lag luam tau txais.
5. Dej nag thiab ntxuav:
Tom qab kev kho mob, cov tshuaj hydrolyzed yog kho nrog acid kom precipitate lub hom phiaj cov khoom. Tom qab ntawd, cov pellet tau muab tshem tawm hauv dej, ua raws li kev ntxuav hnyav kom tshem tawm impurities.
6. Purification:
Cov kauj ruam kawg yog kev ua kom huv ntawm cov khoom xav tau, feem ntau yog siv cov kua chromatography siab. Thaum lub sij hawm tus txheej txheem no, lub purity ntawm cov khoom yuav txiav txim tau los ntawm kev soj ntsuam lub ncov ntawm cov tshuaj nyob rau hauv lub loj spectrum. Nyob rau hauv luv luv, cov tshuaj synthesis ntawm GLP -1 yuav tsum tau ntau rounds ntawm complex tshuaj tiv thaiv thiab nruj purification txheej txheem kom thaum kawg tau txais lub hom phiaj cov khoom.
Txoj kev 4, biosynthesis:
GLP-1 yog ib qho tseem ceeb polypeptide cov tshuaj hormones nrog ntau yam physiological cuam tshuam, nrog rau kev txhawb nqa insulin secretion, suppressing qab los noj mov, txo lub cev hnyav thiab tswj cov insulin rhiab heev, thiab lwm yam. Cov txheej txheem biosynthesis ntawm GLP-1 feem ntau yog tsim los ntawm L hlwb. nyob rau hauv lub caj pas pancreatic, thiab nws cov synthesis tus nqi yog tswj nrog kev noj haus. Cov kauj ruam ntxaws tau qhia raws li hauv qab no:
1. Npaj ua hauj lwm ua ntej synthesis:
Ua ntej biosynthesis ntawm GLP-1, qee qhov kev npaj yuav tsum tau ua, suav nrog kev txiav txim siab hom cell siv, teeb tsa kab lis kev cai thiab xaiv qhov tsim nyog catalytic enzyme. L hlwb yog lub hauv paus tseem ceeb ntawm GLP-1 synthesis vim tias lawv muaj ob hom tshuaj hormones, GIP (glucagon-zoo li peptide 1) thiab GLP-1. L hlwb tuaj yeem raug cais tawm ntawm txoj hnyuv epithelium ntawm luav lossis nas. Ua ntej biosynthesis, hlwb yuav tsum tau coj mus rau ib tug txaus tus naj npawb, thiab txaus cov as-ham thiab tsim kab lis kev cai tej yam kev mob yuav tsum tau muab. Tsis tas li ntawd, nws yog ib qho tsim nyog los xaiv qhov tsim nyog catalytic enzyme los txhawb cov tshuaj tiv thaiv.
2. Synthesis thiab processing ntawm precursors:
Lub biosynthesis ntawm GLP-1 feem ntau tshwm sim hauv L hlwb, thiab nws cov precursor yog tsim los ntawm ob cov tshuaj hormones, GIP thiab GLP-1. Tom qab nkag mus rau hauv cov hlwb endocrine, GIP thiab GLP-1 tau ua tiav los ntawm cov enzymes proteolytic thiab cleaved rau hauv ib tus neeg peptides. Cov enzymes thiab cofactors tau koom nrog hauv cov txheej txheem no, suav nrog precursor polypeptide acidase (PC2), isomerase, thiab lig adhesion yam.
3. Kev sib koom ua ke ntawm polypeptide ntu:
Tom qab ua tiav, GIP thiab GLP-1 peptides tau rov ua dua los ua GLP-1 polypeptide. Cov txheej txheem no yuav tsum tau siv cov glucagon-zoo li peptide 1 (GLP-1) ua tus qauv uas lwm tus neeg peptides tau ua ke los tsim cov polypeptides tshiab. Cov txheej txheem no tseem xav tau qee cov enzymes tshwj xeeb thiab yam tseem ceeb, suav nrog Prohormone Convertase 1/3 (PC1/3) thiab Carboxypeptidase E (CPE).
4. GLP-1 zais cia:
Tom qab GLP-1 yog synthesized thiab ua tiav, nws tau muab cia rau hauv cytoplasm thiab vesicles ntawm endocrine hlwb. Thaum txhawb los ntawm kev noj zaub mov, cov hlwb endocrine tso GLP-1 thiab nkag mus rau cov ntshav ncig los ntawm microvessels. Cov txheej txheem no yog tswj hwm thiab tswj los ntawm ntau txoj hauv kev ntawm cov teeb liab hloov pauv, suav nrog cAMP-Ca2 plusthiab lwm yam.
Hauv luv luv, biosynthesis ntawm GLP-1 cuam tshuam nrog kev sib koom ua ke ntawm ntau qhov txuas thiab ntau yam. Kev sib xyaw ntawm biosynthesis thiab tshuaj synthesis tuaj yeem muab lub hauv paus zoo dua thiab kev txhawb nqa rau kev tshawb fawb thiab tsim khoom ntawm GLP-1.
Txoj kev 5, enzymatic synthesis:
Enzymatic synthesis yog cov synthesis ntawm peptide chains los ntawm catalysis ntawm biological enzymes. Piv nrog rau cov txheej txheem ua kua-theem synthesis ib txwm muaj, enzymatic synthesis tuaj yeem nqa tawm ntawm chav tsev kub, thiab ntau yam khoom siv raw tuaj yeem xaiv. Enzymes xws li theta-kua synthase, AEP, ACE, thiab lwm yam. feem ntau yog siv los ua kom cov synthesis.
Hauv kev xaus, cov kev hais saum toj no yog txoj hauv kev ua tau rau GLP-1 synthesis. Cov txheej txheem sib txawv yog tsim rau cov kev sim sib txawv thiab cov chaw tsim tshuaj.