Single-enzyme kinetics of CALB-catalyzed hydrolysis

被引：155

作者：

Velonia, K ^{[1
]}

Flomenbom, O

Loos, D

Masuo, S

Cotlet, M

Engelborghs, Y

Hofkens, J

Rowan, AE

Klafter, J

Nolte, RJM

de Schryver, FC

机构：

[1] Raboud Univ Nijmegen, Dept Organ Chem, Inst Mol & Mat, NL-6525 ED Nijmegen, Netherlands

[2] Katholieke Univ Leuven, Dept Chem, B-3001 Heverlee, Belgium

[3] Tel Aviv Univ, Sackler Fac Exact Sci, Sch Chem, IL-69978 Tel Aviv, Israel

[4] Dept Biochem, Lab Chem & Biol Dynam, B-3001 Heverlee, Belgium

来源：

ANGEWANDTE CHEMIE-INTERNATIONAL EDITION | 2005年 / 44卷 / 04期

关键词：

analytical methods; enzymes; fluorescence; microscopy; kinetics; single-molecule studies;

D O I：

10.1002/anie.200460625

中图分类号：

O6 [化学];

学科分类号：

0703 ;

摘要：

Real-time measurement of the catalysis and substrate kinetics of a single-enzyme hydrolysis reaction is demonstrated with confocal fluorescence microscopy (CFM; see picture, green = CFM beam). A single lipase is shown to have a broad range of conformations; each conformation contributes to the overall enzymatic activity, an observation that is often masked by ensemble measurements.

引用

页码：560 / 564

页数：5

共 27 条

[1]

Abramowitz M., 1964, HDB MATH FUNCTIONS F

[2] One biocatalyst - Many applications: The use of Candida antarctica B-lipase in organic synthesis [J].

Anderson, EM ;

Karin, M ;

Kirk, O .

BIOCATALYSIS AND BIOTRANSFORMATION, 1998, 16 (03) :181-204

[3] Probing single-molecule T4 lysozyme conformational dynamics by intramolecular fluorescence energy transfer [J].

Chen, Y ;

Hu, DH ;

Vorpagel, ER ;

Lu, HP .

JOURNAL OF PHYSICAL CHEMISTRY B, 2003, 107 (31) :7947-7956

[4] The fluctuating enzyme:: a single molecule approach [J].

Edman, L ;

Földes-Papp, Z ;

Wennmalm, S ;

Rigler, R .

CHEMICAL PHYSICS, 1999, 247 (01) :11-22

[5] Memory landscapes of single-enzyme molecules [J].

Edman, L ;

Rigler, R .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2000, 97 (15) :8266-8271

[6]

FLOMENBOM O, IN PRESS P NATL ACAD

[7] Biological physics [J].

Frauenfelder, H ;

Wolynes, PG ;

Austin, RH .

REVIEWS OF MODERN PHYSICS, 1999, 71 (02) :S419-S430

[8] The role of structure, energy landscape, dynamics, and allostery in the enzymatic function of myoglobin [J].

Frauenfelder, H ;

McMahon, BH ;

Austin, RH ;

Chu, K ;

Groves, JT .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2001, 98 (05) :2370-2374

[9] IMAGING OF SINGLE FLUORESCENT MOLECULES AND INDIVIDUAL ATP TURNOVERS BY SINGLE MYOSIN MOLECULES IN AQUEOUS-SOLUTION [J].

FUNATSU, T ;

HARADA, Y ;

TOKUNAGA, M ;

SAITO, K ;

YANAGIDA, T .

NATURE, 1995, 374 (6522) :555-559

[10] Single-molecule fluorescence spectroscopy of enzyme conformational dynamics and cleavage mechanism [J].

Ha, TJ ;

Ting, AY ;

Liang, J ;

Caldwell, WB ;

Deniz, AA ;

Chemla, DS ;

Schultz, PG ;

Weiss, S .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1999, 96 (03) :893-898

← 1 2 3 →