FEBS Letters | |
Inactivation of human thrombin by water‐soluble carbodiimides | |
Miner, Lydia A.1  Borders, C.L.1  Weerasuriya, Yohan M.1  Chan, Vivien W.F.1  | |
[1] Department of Chemistry, The College of Wooster, Wooster, OH 44691, USA | |
关键词: Thrombin; Carboxyl; essential; Carbodiimide; EDC; EAC; DAPA; EDC; 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride; EAC; 1-ethyl-3-(4-azonia-4; 4-dimethylpentyl)carboiimide iodide; DAPA; dansyl-L-arginine N-(3-ethyl-1; 5-pentanediyl)-amide; Mes; 2-(N-morpholino)ethanesulfonic acid; Hepes; N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid; S-2238; tosyl-D-phenylalanyl-D-pipecolyl-L-arginine-p-nitroanilide; k′; the pseudo-first order rate constant for inactivation of thrombin by ECD or EAC under a given set of conditions; K 50; the DAPA concentration at which thrombin is half-saturated; | |
DOI : 10.1016/0014-5793(89)81123-8 | |
学科分类:生物化学/生物物理 | |
来源: John Wiley & Sons Ltd. | |
【 摘 要 】
Human thrombin is inactivated by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) or 1-ethyl-3-(4-azonia-4,4-dimethylpentyl)carbodiimide (EAC) by a second order process. A plot of the pseudo-first order rate constant for inactivation by 20 mM EDC at different pH values from 4.2 to 7.7 at 25°C shows that this inactivation is due to the modification of a protonated carboxyl with a pK a of 5.6. The rate of inactivation by EDC at pH 6.0 is reduced, but not eliminated by saturating levels of the competitive inhibitor dansyl-L-arginine N-(3-ethyl-1,5-pentanediyl)amide, suggesting that the essential carboxyl modified is one other than Asp-189 in the substrate specificity pocket of thrombin. Complete inactivation by 14C-EAC correlates with the incorporation of approximately 2.5–3 EACs per thrombin.
【 授权许可】
Unknown
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