Zogulitsa

Kumvetsetsa kagayidwe kachakudya kamene kamakhala kofunikira kuti pakhale chitukuko ndi kukhathamiritsa kwa ma cell athunthu a biocatalytic, chifukwa kumapangitsa kupanga bwino.Izi ndizowona makamaka pa redox biocatalysis pomwe ma cell a metabolic omwe akugwira ntchito amagwiritsidwa ntchito chifukwa cha cofactor/cosubstrate regenerative capacity yokhazikika mwa wolandirayo.Recombinant Escherichia coli inagwiritsidwa ntchito popanga proline-4-hydroxylase (P4H), dioxygenase yomwe imayambitsa hydroxylation ya L-proline yaulere mu trans-4-hydroxy-L-proline ndi-ketoglutarate (a-KG) monga cosubstrate.Mu biocatalyst ya cell yonseyi, kagayidwe wapakati wa kaboni umapereka cosubstrate a-KG yofunikira, kuphatikiza P4H biocatalytic performance molunjika ku carbon metabolism ndi metabolism.Pogwiritsa ntchito zida zonse zoyesera komanso zowerengera za biology, monga metabolic engineering ndi (13)C-metabolic flux analysis ((13)C-MFA), tinafufuza ndi kulongosola mochulukira kuyankha kwa thupi, kagayidwe kachakudya, ndi mphamvu ya bioenergetic ya cell cell biocatalyst. kupita ku bioconversion yomwe imayang'aniridwa ndikuzindikira zovuta za kagayidwe kachakudya kuti mupititse patsogolo luso la uinjiniya wa njira.Kusintha kwa ma cell athunthu okhala ndi mtundu wosinthika woterewu sikunangoyambitsa kuchuluka kwa proline hydroxylation komanso kuwirikiza kawiri kwa kuchuluka kwa mapangidwe a trans-4-L-hydroxyproline (hyp), poyerekeza ndi mtundu wakuthengo.Kuwunika kwa carbon flux kudzera pakatikati pa metabolism ya mutant strain kunawonetsa kuti kuwonjezeka kwa KG kwa ntchito ya P4H sikunapititse patsogolo kutulutsa kwa-KG, kusonyeza kuti TCA imagwira ntchito mokhazikika motsatira momwe amaphunzirira.Pazovuta zamtundu wakuthengo, kaphatikizidwe ka P4H ndi catalysis zidapangitsa kuchepa kwa zokolola za biomass.Chochititsa chidwi n'chakuti, ΔputA strain inawonjezeranso kutayika kwa ATP ndi NADH pochepetsa kufunidwa kwa mphamvu zosamalira pa mlingo wochepa wa shuga, m'malo mowonjezera ntchito ya TCA. The putA knockout in recombinant E. coli BL21(DE3)(pLysS) inapezeka kukhala akulonjeza kuti apange P4H catalysis yopindulitsa osati ponena za zokolola za biotransformation, komanso zokhudzana ndi mitengo ya biotransformation ndi proline uptake ndi zokolola za hyp pa gwero la mphamvu.Zotsatira zikuwonetsa kuti, pakugogoda kwa putA, kuphatikiza kwa TCA-cycle kuti proline hydroxylation kudzera pa cosubstrate a-KG kumakhala chinthu chofunikira kwambiri komanso chandamale chopititsira patsogolo luso la ma biotransformations omwe amadalira a-KG.