Ukwahlula kunye nokuhlanjululwa kweeprotheni zisetyenziswa ngokubanzi kuphando lwe-biochemistry kunye nokusetyenziswa kwaye sisakhono esibalulekileyo sokusebenza. Iseli eqhelekileyo ye-eukaryotic inokuqulatha amawaka eeprotheyini ezahlukeneyo, ezinye zizityebi kakhulu kwaye ezinye zineekopi ezimbalwa kuphela. Ukuze ufunde okuthileiprotheni, kuyimfuneko ukuqala ukuhlambulula iprotheni evela kwezinye iiprotheni kunye ne-non-protein molecules.

1. Indlela yokukhupha ityuwaiprotheni:

Ityuwa engathathi hlangothi inempembelelo ebalulekileyo kwi-solubility yeprotheni. Ngokuqhelekileyo, ngokunyuka koxinzelelo lwetyuwa phantsi koxinzelelo lwetyuwa ephantsi, ukunyibilika kweprotheni kuyanda. Oku kubizwa ngokuba yityuwa; xa ityuwa iqhubeka ikhula, I-solubility yeprotheni iyancipha ukuya kumaqondo ahlukeneyo kwaye ihlukanise enye emva kwenye. Le nto ibizwa ngokuba yi-salting out.

2. Indlela yokupakisha inqaku le-isoelectric:

I-electrostatic repulsion phakathi kwamasuntswana yeyona incinci xa iprotein imile, ngoko ke ukunyibilika kolona kuncinci. Amanqaku e-isoelectric eeprotheni ezahlukeneyo ahlukile. I-pH yesisombululo se-conditioning ingasetyenziselwa ukufikelela kwindawo ye-isoelectric yeprotheyini Yenze ukuba iqokelele, kodwa le ndlela ayifane isetyenziswe yodwa kwaye ingadibaniswa kunye nendlela yokukhupha i-salting.

3.I-Dialysis kunye ne-ultrafiltration:

I-Dialysis isebenzisa i-membrane ene-semi-permeable ukwahlula iiprotheni ezinobungakanani obuhlukeneyo bemolekyuli. Indlela ye-ultrafiltration isebenzisa uxinzelelo oluphezulu okanye amandla e-centrifugal ukwenza amanzi kunye nezinye iimolekyuli ezincinci ze-solute zidlule kwi-membrane evumelekileyo, ngelixaiprotheniihlala kwi-membrane. Unokukhetha ubungakanani obuhlukeneyo bepore ukubamba iiproteni zobunzima bemolekyuli ezahlukeneyo.

Indlela yokuhluza ye-4.Gel:

Ikwabizwa ngokuba yi-chromatography yobungakanani okanye i-molecular sieve chromatography, le yenye yezona ndlela ziluncedo zokwahlula imixube yeeproteni ngokobungakanani bemolekyuli. Izinto ezisetyenziswa ngokuqhelekileyo ukupakisha kwikholamu i-glucose gel (Sephadex ged) kunye ne-agarose gel (i-agarose gel).

5.I-Electrophoresis:

Ngaphantsi kwemeko efanayo ye-pH, iiprotheyini ezahlukeneyo zinokwahlulwa ngenxa yobunzima bemolekyuli ezahlukeneyo kunye neentlawulo ezahlukeneyo kwintsimi yombane. Kufanelekile ukunikela ingqalelo kwi-isoelectric set electrophoresis, esebenzisa i-ampholyte njengomthwali. Ngexesha le-electrophoresis, i-ampholyte yenza i-pH gradient ngokuthe ngcembe idityaniswe ukusuka kwi-electrode elungileyo ukuya kwi-electrode engalunganga. Xa iprotheni enentlawulo ethile ibhukuda kuyo, iya kufikelela omnye komnye. Indawo ye-pH yendawo yombane iyayeka, kwaye le ndlela ingasetyenziselwa ukuhlalutya nokulungiselela iiprotheni ezahlukeneyo.

6.Ion unxibelelwano lwechromatography:

Ii-agent zonxibelelwano ze-ion ziquka ii-agent zonxibelelwano ze-cationic (ezifana ne-carboxymethyl cellulose; i-CM-cellulose) kunye nee-agent zonxibelelwano ze-anionic (i-diethylaminoethyl cellulose). Xa udlula kwikholamu yonxibelelwano lwe-ion chromatography, iprotheyini enentlawulo echaseneyo ne-agent yonxibelelwano ye-ion ibhengezwa kwi-agent yonxibelelwano ye-ion, kwaye emva koko i-adsorbed.iprotheniihlanjululwa ngokutshintsha i-pH okanye amandla e-ionic.

7. Ichromatography ehambelanayo:

I-Affinity chromatography yeyona ndlela iluncedo kakhulu yokwahlula iiproteni. Kuhlala kufuna inyathelo elinye kuphela ukwahlula iprotheyini ethile ukuze ihlanjululwe kumxube weprotheyini emdaka kunye nokucoceka okuphezulu.

Le ndlela isekelwe kwi-covalent yokubophelela kwiiprotheni ezithile kwenye i-molecule ebizwa ngokuba yi-ligand (Ligand).

Umgaqo osisiseko:

iiproteni zikhona kumxube omdaka kwizicubu okanye iiseli, kwaye uhlobo ngalunye lweseli lunamawaka eeproteni ezahlukeneyo. Ngoko ke, ukuhlukana phakathi kweeprotheni yinxalenye ebalulekileyo ye-biochemistry, kwaye ayizange ibe yodwa. Okanye isethi yeendlela esele zenziwe ziyakwazi ukususa naluphi na uhlobo lweprotheyini kwiprotheyini edibeneyo edibeneyo, ngoko ke iindlela ezininzi zihlala zisetyenziswa ngokudibeneyo.