University of North Florida
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Stuart Chalk, Ph.D.
Department of Chemistry
University of North Florida
Phone: 1-904-620-1938
Fax: 1-904-620-3535
Email: schalk@unf.edu
Website: @unf

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Kidney

Classification: Biological tissue -> kidney -> hog

Citations 2

"Enzyme Assay By Repetitive Flow Injection Analysis. Application To The Assay Of Hog Kidney Aminoacylase"
Anal. Lett. 1984 Volume 17, Issue 7 Pages 589-598
John W. Keller

Abstract: The initial rate of methionine release from N-acetyl-L-methionine by the cited enzyme was measured by injecting portions of the assay solution into a stream of 0.1 M borate buffer of pH 9.7 that contained 0.0083% (v/v) of phthalaldehyde and 0.17 mg mL-1 of 2-mercaptoethanol; the reaction product was detected spectrophotometrically at 336 nm. The flow injection system comprised a Rheodyne type 50 four-way PTFE rotary valve, with 0.8-mm flow passages and a 70 µL sample loop, connected in series to a coiled PTFE tube (50 cm x 0.5 mm) and a 1-cm-pathlength flow-through cell (40 µL). The buffer solution was pumped at 1 mL min-1. Sample was injected every 1.5 min after initiation of the enzyme reaction, and 10 to 15 injections were made over a 15 to 22-min period. The sensitivity could be improved by 20% by use of a stopped-flow technique. Activities measured in this way agreed with those by a manual initial-rate ninhydrin assay.
Aminoacylase Methionine Clinical analysis Spectrophotometry Stopped-flow Enzyme

"Aminoacylase I From Hog Kidney: Anion Effects And The PH Dependence Of Kinetic Parameters"
Biochim. Biophys. Acta 1988 Volume 959, Issue 3 Pages 370-377
Johannes Henseling and Klaus-Heinrich Röhm*

Abstract: The hydrolysis of acetylamino acids by highly purified hog kidney aminoacylase I (N-acylamino acid amidohydrolase, EC 3.5.1.14) was investigated using flow injection analysis to determine reaction rates. We show that the distinctly bell-shaped pH versus activity profiles observed in previous studies do not reflect protonic equilibria in the enzyme, but were created by buffer effects. At low pH, anions such as phosphate, nitrate or chloride markedly increase Km. These effects are reversed at higher pH. In zwitterionic 'Good' buffers (Mes, Mops, and Bicine), maximal velocities are almost independent of pH between 6.5 and 9 for all substrates studied (Ac-LAla, Ac-LGlu, Ac-LMet, Ac-LPhe). Below pH 6.5, the catalytic constants decrease with pH, apparently due to the protonation of a carboxylate with a pKa of 5.5-6. The pH dependence of Km markedly varies among different substates. We conclude that the observed profiles all result from the dissociation of an active-site residue with a pKa of 8-8.5, which we tentatively identify as an active-site cysteine residue. A working model of aminoacylase catalysis is presented that accounts for most of the known facts.
Amino acids, acetyl Catalysis Kinetic Optimization