A Systematic Approach to Kinetic Studies of Multisubstrate Enzyme Systems

Author(s):  
James R. Fisher ◽  
Vincent D. Hoagland
1952 ◽  
Vol 30 (3) ◽  
pp. 306-317 ◽  
Author(s):  
N. Hansl ◽  
E. R. Waygood

Spinach leaf carbonic anhydrase has been used as a tool in the Krebs-Roughton technique to determine whether several plant decarboxylase systems give rise to carbon dioxide or bicarbonate as the primary end product. The results show that in addition to the urease – urea and yeast carboxylase – pyruvic systems, the plant enzyme systems decarboxylating pyruvic, oxalacetic, glutamic, and α-ketoglutaric acids produce carbon dioxide and not bicarbonate as the primary end product.


1968 ◽  
Vol 57 (4) ◽  
pp. 623-638 ◽  
Author(s):  
W. Hoffmann ◽  
H. Breuer

ABSTRACT The effect of 1,2α-methylene-6-chloro-pregna-4,6-dien-17α-ol-3,20-dione (Cyproterone) on the formation of C19-steroids from 17α-hydroxyprogesterone was studied in the 20 000 × g supernatants and the microsomal fractions obtained from the testis of young (5 weeks ± 5 days) and old (5 months ± 14 days) rats. Under the experimental conditions chosen, cyproterone (final concentration 2.7-27 μmoles/1) inhibited the biogenesis of C19-steroids to an extent of 20–80%. On the basis of enzyme-kinetic studies, cyproterone belongs to the class of »mixed type inhibitors«. The investigation of the pattern of the C19-steroid metabolites showed that, after the addition of cyproterone, the formation of testosterone, androst-4-ene-3,17-dione and the »polar« fraction was inhibited, whereas the androsterone fraction mostly increased. These results suggest that cyproterone affects not only the activity of the desmolase, but also the activities of the enzymes involved in the metabolism of C19-steroids. The experiments reported are, to a large extent, in agreement to the already known fact that the activity of the desmolase depends upon the age of rats. However, the biogenesis of C19-steroids from C21-steroids is greatly influenced by the preparation of the relevant enzyme systems and by the experimental conditions.


2004 ◽  
Vol 71 ◽  
pp. 1-14
Author(s):  
David Leys ◽  
Jaswir Basran ◽  
François Talfournier ◽  
Kamaldeep K. Chohan ◽  
Andrew W. Munro ◽  
...  

TMADH (trimethylamine dehydrogenase) is a complex iron-sulphur flavoprotein that forms a soluble electron-transfer complex with ETF (electron-transferring flavoprotein). The mechanism of electron transfer between TMADH and ETF has been studied using stopped-flow kinetic and mutagenesis methods, and more recently by X-ray crystallography. Potentiometric methods have also been used to identify key residues involved in the stabilization of the flavin radical semiquinone species in ETF. These studies have demonstrated a key role for 'conformational sampling' in the electron-transfer complex, facilitated by two-site contact of ETF with TMADH. Exploration of three-dimensional space in the complex allows the FAD of ETF to find conformations compatible with enhanced electronic coupling with the 4Fe-4S centre of TMADH. This mechanism of electron transfer provides for a more robust and accessible design principle for interprotein electron transfer compared with simpler models that invoke the collision of redox partners followed by electron transfer. The structure of the TMADH-ETF complex confirms the role of key residues in electron transfer and molecular assembly, originally suggested from detailed kinetic studies in wild-type and mutant complexes, and from molecular modelling.


2017 ◽  
Vol 2 (1) ◽  
pp. 86-94 ◽  
Author(s):  
Lindsay Heggie ◽  
Lesly Wade-Woolley

Students with persistent reading difficulties are often especially challenged by multisyllabic words; they tend to have neither a systematic approach for reading these words nor the confidence to persevere (Archer, Gleason, & Vachon, 2003; Carlisle & Katz, 2006; Moats, 1998). This challenge is magnified by the fact that the vast majority of English words are multisyllabic and constitute an increasingly large proportion of the words in elementary school texts beginning as early as grade 3 (Hiebert, Martin, & Menon, 2005; Kerns et al., 2016). Multisyllabic words are more difficult to read simply because they are long, posing challenges for working memory capacity. In addition, syllable boundaries, word stress, vowel pronunciation ambiguities, less predictable grapheme-phoneme correspondences, and morphological complexity all contribute to long words' difficulty. Research suggests that explicit instruction in both syllabification and morphological knowledge improve poor readers' multisyllabic word reading accuracy; several examples of instructional programs involving one or both of these elements are provided.


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