resonance (REDOR) and cross-polarization magic-angle spinning (CPMAS)-echo NMR experiments to be performed. After a long prelyophilization period, the gradual formaPlants and microorganisms contain an en31 13 zyme called EPSP synthase, which catalyzes tion of a strong P- C coupling, consistent with a tetrahedral intermediate is observed. the formation of inorganic phosphate and EPSP, a precursor to aromatic amino acids. Shorter low-temperature incubation periods resulted in spectra of previously uncharacGlyphosate, an herbicide that is nontoxic to terized intermediates. mammals, forms a strong complex with the enzyme, inhibiting the Based on observaformation of EPSP in tions of the forward plants. Daniel R. Studelreaction, the study supska and co-workers at ports the formation of Washington University an enzyme-PEP comreport a new method for plex, followed by redetermining intermediate moval of phosphate to MPE fluorescence detector for CE steps involved in EPSP yield a covalently separations. A: Microscope objective synthase reaction pathbound enolpyruvylfocusing near-1R laser at capillary outlet. B: ways via solid-state magicDetection scheme showing beam splitters enzyme adduct. Upon (B), dichroic mirrors (D), filters (F), and angle spinning NMR addition of S3P-OH to photomultiplier tubes (PM). the adduct, an enzymeThe method involves ketal intermediate is subzero entrapment of observed which underCE combined with MPE fluorescence the substrates shikimate detection is a sensitive and versatile ap3-phosphate (S3P) and cleavage to give proach that could prove useful for singlephosphoenolpyruvate EPSP. The authors becell analyses. Correlation of the fluores(PEP) in EPSP synthase, lieve that the wellcence of peptides, proteins, and monoamfollowed by lyophilization characterized tetraheine neurotransmitters to levels of redox with stabilizers. The fordral intermediate cofactors in a cell could provide informamation of EPSP is tempobe formed only after tion regarding processes such as energy rarily halted by cooling EPSP is generated metabolism and neurotransmitter regulathe sample, allowing (Biochemistry y997 Reaction scheme for the formation tion. (J. Am. Chem. Soc. 1998,120,18-22) rotational-echo double36 15555-60) of EPSP.
EPSP intermediates by NMR . . .
. . . and EPSP intermediates by rapid mixing/ESIMS
the well-characterized reaction involving the enzyme EPSP synthase and its catalytic role in the formation of EPSP. The reaction had been previously shown to proceed through a tetrahedral intermediate. Results Chemical quenching methods are comconsistent with a tetrahedral intermediate monly used to identify enzyme reaction inare obtained by the rapid-mixing/ESIMS termediates; however, some intermediates are too labile under quenching conditions to technique without chemical quenching. be detected and characterized by such methThe extent to which a reaction proceeds ods. Lee Q. Huang, Karen S. Anderson, ,nd is controlled by the length of the capillary co-workers at Bayer column and the mixCorporation and ing speed. The reacYale University tion time is varied by School of Medicine allowing the soludescribe a new aptions to travel proach for the dethrough the column tection of enzyme together, at different intermediates on a rates. The technique subsecond time provides a way to scale with a "pulsedobserve transient flow" rapid-mixing enzyme intermedidevice and electroates that are too laionization MS bile to be detected (ESIMS) by quenching methods (Biochemistry The method is Illustration of a rapid-mix, pulsed-flow ESI 1997 36 15472-76) demonstrated on system.
Probing the oil/water interface Surfactants play an important role in everyday life. They're found in motor oils, lubricants, detergents, and soaps. Despite their widespread use, little is known about how water and surfactants interact at liquid surfaces and interfaces. G. L. Richmond and D. E. Gragson of the University of Oregon use vibrational sum-frequency generation to obtain vibrational spectra of water molecules at an oil/water interface in the presence of a charged, soluble surfactant. In this study, they use an optical parametric amplifier system that produces IR pulses tunable over the range 2.4-4.0 um. The IR pulses are combined at the oil/water interface with 800-nm light from a titaniumsapphire regenerative amplifier. Spectra from the oil/water interface were obtained in an internal reflection geometry, with each beam incident at the critical angle for that wavelength. Spectra were taken as the bulk concentration of sodium dodecyl sulfate (SDS) and the ratio of H20 to D20 were varied,
Analytical Chemistry News & Features, March 1, 1998 173 A
