Development of a microdroplet mixing technique for the study of rapid reactions by Raman spectroscopy

Simpson, Stanley F.; Kincaid, James R.; Holler, F. James

doi:10.1021/ac00127a054

Cited by 25 publications

(11 citation statements)

References 35 publications

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“…A 394 /A 280 ) Ն 3.5, was used for spectroscopic measurement immediately after preparation. The Compound I species are prepared by using compressed nitrogen to force the enzyme and buffered hydrogen peroxide solutions (Aldrich) (enzyme:H 2 O 2 ϭ 1:2.75), held in separate reservoirs, into a mixing chamber and subsequently through the microdroplet generator (30,31). A low temperature stirred cell (36) was employed for the measurement of Compound II species, while a conventional spinning cell was used for the resting state samples.…”

Section: Methodsmentioning

confidence: 99%

“…Later studies, using conventional flow methods (25) or short laser pulses (26,27), yielded spectra which were insignificantly different from that of HRP-II and were interpreted as being either that of HRP-I (25) or that of an HRP-II-like photoproduct (27). We (28) and others (29) attempted to overcome this photolability problem by using a novel microdroplet sampling device, first employed in our laboratory (28,30,31), which effectively reduces sample residence time in the beam (to less than 5 s). The results of these latter two studies were in essential agreement, giving a new spectrum at low laser powers which, at higher powers, reverted to the spectrum of a (presumably photoreduced) product which resembled that of authentic HRP-II.…”

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confidence: 99%

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Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Kincaid

Zheng

Al-Mustafa

et al. 1996

Journal of Biological Chemistry

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Resonance Raman studies of native and mesohemereconstituted horseradish peroxidase and their catalytic intermediates, known as Compounds I and II, have been conducted using both near UV (ϳ350 nm) and visible (406.7 nm) excitation. Careful power studies indicate that the authentic Compound I spectra are obtainable using near UV excitation, but that use of visible excitation results in contamination of the Compound I spectrum with the spectrum of a Compound II-like photoproduct. Using H 2 18 O 2 , the (Fe‫؍‬O) stretching modes for both systems are unambiguously identified, for the first time, at ϳ790 cm ؊1 . The authentic Compound I spectra are indicative of an 2 A 1u -like ground state for both the native and the mesoheme-reconstituted proteins. Finally, the possible biological implications of such information are briefly discussed.

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Section: Methodsmentioning

confidence: 99%

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confidence: 99%

Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Kincaid

Zheng

Al-Mustafa

et al. 1996

Journal of Biological Chemistry

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“…The age of the sample at the laser spot is calculated from the flow rate to be approximately 260 and 652 ms for the two flow rates employed (1.84 and 4.6 mL/min). The mixing efficiency was tested by monitoring the reaction of HSO; with OH-(as described by Simpson et al, 1986); the diffusion-controlled reaction was found to be essentially complete under the range of flow rates used for the experiment.…”

Section: Formation Of the Dienolatementioning

confidence: 99%

Substrate polarization by residues in Δ⁵‐3‐ketosteroid isomerase probed by site‐directed mutagenesis and UV resonance Raman spectroscopy

Austin

Kuliopulos

et al. 1992

Protein Science

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A5-3-Ketosteroid isomerase (KSI: EC 5.3.3.1) of Pseudomonas testosteroni catalyzes the isomerization of A5-3-ketosteroids to A4-3-ketosteroids by the stereospecific transfer of the steroid 40-proton to the 60-position, using Tyr-14 as a general acid and Asp-38 as a base. Ultraviolet resonance Raman (UVRR) spectra have been obtained for the catalytically active double mutant Y55F + Y88F, which retains Tyr-14 as the only tyrosine residue (referred to as the Y140 mutant), and the Y14F mutant, which has 50,000-fold lower activity. The UVRR results establish that binding of the product analog and competitive inhibitors 19-nortestosterone or 4-fluoro-19-nortestosterone to the Y140 mutant does not result in the formation of deprotonated Tyr-14. The UVRR spectra of the steroid inhibitors show large decreases in the vinyl and carbonyl stretching frequencies on binding to the Y140 enzyme but not on binding to the Y14F enzyme. These changes cannot be mimicked by protonation of the steroids. For 19-nortestosterone, the vinyl and carbonyl stretching frequencies shift down (with respect to the values in aqueous solution) by 18 and 27 cm", respectively, on binding to Y140 KSI. It is proposed that the changes in the steroid resonance Raman spectrum arise from polarization of the enone moiety via the close proximity of the charged Asp-38 side chain to the vinyl group and the directional hydrogen bond between Tyr-14 and the 3-carbonyl oxygen of the steroid enone. The 230-nm-excited UVRR spectra do not, however, show changes that are characteristic of strong hydrogen bonding from the tyrosine hydrogen. It is proposed that this hydrogen bonding is compensated by a second hydrogen bond to the Tyr-14 oxygen from another protein residue.UVRR spectra of the Y14, enzyme obtained using 200 nm excitation show enhancement of the amide I1 and S Raman bands. The secondary structure of KSI was estimated from the amide I1 and S intensities and was found to be low in a-helical structure. The a-helix content was estimated to be in the range of 0-25% (i.e., 10 15%).

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“…A spatially and temporally reproducible generation of the liquid–liquid interface is essential to conduct investigations on the miscible interface. In that sense, liquid droplets facilitate the studies on the dynamics at the liquid–liquid interface, because small liquid droplets can be generated reproducibly by the use of a piezo droplet nozzle. , Simpson and co-workers have applied the collision of liquid droplets for the investigation of the rapid reaction kinetics triggered by mixing two solutions. − They merged two streams of droplets and observed the amount of the reactant and product species by Raman spectroscopy. They determined the rate constant of the complex-formation reaction of Fe 2+ and 1,10-phenanthroline by using this method .…”

Section: Introductionmentioning

confidence: 99%

Initial Collision Process of Two Miscible Droplets

Anahara

Kohno

2017

J. Phys. Chem. B

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Dynamic properties of the metastable interface between two miscible solutions are investigated by the collision of two droplets. A clear interface is observed between the two colliding droplets. The interface moves in the colliding droplet toward the side where the original droplet has a lower surface tension. The interface is set to the middle of the colliding droplet by controlling the surface tension of the droplets to observe the chemical reactions at the droplet interface by cavity-enhanced Raman spectroscopy. This study provides a foundation for further research on the initial process of the chemical reactions of two miscible solutions.

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Development of a microdroplet mixing technique for the study of rapid reactions by Raman spectroscopy

Cited by 25 publications

References 35 publications

Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Substrate polarization by residues in Δ⁵‐3‐ketosteroid isomerase probed by site‐directed mutagenesis and UV resonance Raman spectroscopy

Initial Collision Process of Two Miscible Droplets

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Development of a microdroplet mixing technique for the study of rapid reactions by Raman spectroscopy

Cited by 25 publications

References 35 publications

Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Resonance Raman Spectra of Native and Mesoheme-reconstituted Horseradish Peroxidase and Their Catalytic Intermediates

Substrate polarization by residues in Δ5‐3‐ketosteroid isomerase probed by site‐directed mutagenesis and UV resonance Raman spectroscopy

Initial Collision Process of Two Miscible Droplets

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Substrate polarization by residues in Δ⁵‐3‐ketosteroid isomerase probed by site‐directed mutagenesis and UV resonance Raman spectroscopy