| AUTHOR |
TITLE |
Robert S. Phillips,
Bakthavatsalam Sundararaju,
Nicolai G. Faleev |
Proton Transfer and Carbon-Carbon Bond Cleavage in the Elimination of Indole Catalyzed by Escherichia coli Tryptophan Indole-Lyase
Journal of the American Chemical Society; 2000; ASAP Article |
Experimental Section
Kinetic Measurements. Single-wavelength stopped-flow kinetic measurements and rapid-scanning experiments were performed using an RSM spectrophotometer and a stopped-flow compartment with a 10 mm path length observation cell from OLIS, Inc. This instrument has a mixing dead time of about 2 ms; thus, a reaction with a rate constant of 350 s-1 will lose about half its amplitude during mixing. Rapid-scanning measurements were performed over the range from 325 to 550 nm at 1 kHz, while single-wavelength measurements for the concentration dependencies were collected at 4 kHz. Some of the single-wavelength measurements (Figure 4) were performed on a Kinetics Instruments stopped-flow mixer with a modified Cary 14 UV/vis spectrophotometer (OLIS), and preliminary rapid-scanning experiments were performed with a diode array detector from EG&G Princeton Applied Research, as previously described. |
J. Richard Miller,
Dale E. Edmondson |
Structure-Activity Relationships in the Oxidation of Para-Substituted Benzylamine Analogues by Recombinant Human Liver Monoamine Oxidase A
Biochemistry; 1999; 38(41); 13670-13683. |
| Anaerobic Rapid-Scanning Stopped-Flow Kinetic Experiments with α,α-[1H]-and α,α[2H]-p-Chlorobenzylamine. Conditions and preparation of anaerobic enzyme and substrates were identical to that described for single-wavelength stopped-flow experiments except that 25 mM MAO A was utilized. Experiments were conducted using an OLIS-RSM1000 stopped-flow spectrophotometer (OLIS, Inc., Bogart, GA) in the laboratory of W. David Wilson (Department of Chemistry, Georgia State University, Atlanta, GA). Spectra were acquired across a 150 nm range from 335 to 485 nm. |
Kwang-Hwan Jung,
Elena N. Spudich,
Pervin Dag,
John L. Spudich |
Transducer-Binding and Transducer-Mutations Modulate Photoactive-Site-Deprotonation in Sensory Rhodopsin I
Biochemistry; 1999; 38(40); 13270-13274. |
Flash Photolysis.
Flash-induced absorption changes were recorded by a cross-beam spectrophotometer with a 532 nm, 40 mJ/6 ns pulse Nd:YAG laser (Surelite I, Continuum, Santa Clara, CA) and RSM-1000 spectrophotometer (On-line Instrument System, Inc. Bogart, GA). The absorbance transients due to S373 formation were monitored at 400 nm. Thirty transients were averaged for each trace at 18°C, 25°C, 35°C, and 45°C, and fifty for each 230 nm scan. The t1/2 of S373 deprotonation was calculated by a single-exponential curve-fit using SigmaPlot (Jandel, San Rafael, CA). |
Adele K. Addington,
David A. Johnson |
Inactivation of Human Lung Tryptase: Evidence for a Re-Activatable Tetrameric Intermediate and Active Monomers
Biochemistry; 1996; 35(42); 13511-13518. |
| Intrinsic Fluorescence and Anisotropy Measurements. Fluorescence spectral data were obtained with a Perkin-Elmer 650-40 fluorimeter modified by On-Line Instrument Systems Inc. (Bogart, GA) to provide computer control and data collection with continuous measurement of polarization of fluorescence, incorporating a piezoelectric birefringence modulator and an analyzing polarizer, as described by Wampler and DeSa (1974). |
Chris W. Chronister,
Robert A. Binstead,
Jinfeng Ni,
Thomas J. Meyer |
Mechanism of Water Oxidation Catalyzed by the -Oxo Dimer [(bpy)2(OH2)RuIIIORuIII(OH2)(bpy)2]4+
Inorganic Chemistry; 1997; 36(18); 3814-3815. (Communication) |
| Rapid-scanning, stopped-flow kinetics experiments showed no risetime for the initial spectral change that occurs upon mixing RuIVORuV (pH 6) with acid. |
Vahe Bandarian,
George H. Reed |
Isotope Effects in the Transient Phases of the Reaction Catalyzed by Ethanolamine Ammonia-Lyase: Determination of the Number of Exchangeable Hydrogens in the Enzyme-Cofactor Complex
Biochemistry; 2000; 39(39); 12069-12075. |
| Stopped-Flow Spectrophotometry. An Olis RSM-1000 rapid-scanning stopped-flow spectrophotometer was used for the measurements. Data processing was accomplished using the software package supplied with the instrument. For experiments where the effect of preincubation of EAL with cofactor was examined, EAL (~15 µM sites) and Hepes/NaOH, pH 7.5 (20 mM), were placed in one syringe, and the second syringe was filled with a solution of coenzyme B12 (35 µM), Hepes/NaOH, pH 7.5 (20 mM), and substrate (5 mM). The concentrations of enzyme, cofactor, and substrate were halved after mixing. For all other experiments, one syringe was filled with substrate and buffer as above. EAL, coenzyme B12, and buffer were mixed and placed in the second syringe. All experiments were completed within 10-15 min of mixing of the enzyme with coenzyme B12. Data from at least three repetitions were averaged. All solutions containing coenzyme B12 were handled in dim light. |
Andrea J. Clarkson,
David A. Buckingham,
Andrew J. Rogers,
Allan G. Blackman,
Charles R. Clark |
Kinetic Origin of the Chelate Effect. Base Hydrolysis, H-Exchange Reactivity, and Structures of syn,anti-[Co(cyclen)(NH3)2]3+ and syn,anti-[Co(cyclen)(diamine)]3+ Ions (diamine = H2N(CH2)2NH2, H2N(CH2)3NH2)
Inorganic Chemistry; 2000; ASAP Article. |
Experimental Section
Zero time absorbance data (25.0 °C, I = 1.0 M, NaClO4) were obtained using Durrum D-110 and Olis USA stopped-flow spectrophotometers. Measurements were obtained within 10 ms of mixing equal volumes of the complex in NaClO4 solution and NaOH solution of the required concentration. Rates of alkaline hydrolysis (25.0 °C, I = 1.0 M, NaClO4) were measured using a Cary 219 UV-vis spectrophotometer for [Co(cyclen)en](ClO4)3, and either the Durrum or Olis for [Co(cyclen)tn](ClO4)3, [Co(cyclen)(NH3)2](ClO4)3, and [Co(cyclen)(NH3)OH](ClO4)2. |
