Use of the F test for determining the degree of enzyme-kinetic and ligand-binding data. A Monte Carlo simulation study

1. Initial-rate data were simulated for 13 representative enzyme mechanisms with the use of several distributions of rate constants in order to locate conditions leading to v([S]) curves in physiological ranges of substrate concentration. 2. In all, 420 sets of such v([S]) curves were generated with the use of several choices of substrate concentration range (two, three or four orders of magnitude), number of experimental points (10, 15 or 20), error on v (5-10%) and standard deviation on v (5-9%) in order to simulate experimental results in a number of possible ways. 3. Curve-fitting was carried out to rational functions of degree 1:1, 2:2, . . ., 5:5 until there was no statistically significant decrease in the sum of weighted squared residuals as judged by the F test at 95% and 99% confidence levels. 4. It was checked whether the non-linear regression program had located a good minimum in the sum of squares by also fitting the data with the correct values of parameters as starting estimates. 5. A similar procedure was adopted with 110 sets of binding data simulated for 11 models, and the F test was used to see if fractional-saturation data generated by a binding polynomial of order n could be adequately fitted by one of order m, m less than n. 6. From the 530 simulations the F test was successful in fixing the correct degree with a probability of 0.62 at the 95% confidence level, but this fell with increase in degree as follows: 1:1 (0.98), 2:2 (0.71), 3:3 (0.43) and 4:4 (0.34), the first numbers being the degree of the rate equation and those in parentheses referring to the 95% confidence level. 7. It made little difference whether the 95% or the 99% confidence level was consulted, as there were very few borderline cases. 8. The chance of detecting deviations from Michaelis-Menten kinetics, i.e. terms of at least second-order in a rate equation of degree n:n, n greater than 1, was estimated to be about 0.8. 9. The probability of the F test leading to a spurious result due to error in the data was found to be about 0.04. 10. The probability with which 4:4 mechanisms can lead to v([S]) plots with no, one, two or three turning points was computed, and it was established that there is a small but finite chance that the increase in degree that occurs in some mechanisms when ES in equilibrium EP interconversions are explicitly allowed for can be detected by the F test.     

The sequence GGCmCGG is resistant to MspI cleavage.

MspI essentially fails to cut the sequence GGCmCGG at enzyme concentrations which give total digestion of CCGG, CmCGG and GGCCGG sites. This result explains why certain sites in mammalian DNA are resistant to both MspI and HpaII and shows that this results from an idiosynchracy of MspI rather than a novel form of DNA methylation at this site in mammalian cells.     

VARIATION IN THE DURATION OF MITOSIS IN THE CRYPTS OF LIEBERKUHN OF THE RAT; A CYTOKINETIC STUDY USING VINCRISTINE

The variation in the duration of mitosis ( t _m) with cell position in the small intestinal crypts of the adult rat has been measured by a stathmokinetic technique using vincristine. The value for the whole crypt column was 0.43 hr, or 26 min. At the bottom of the crypt t _m was in excess of 1 hr, but rapidly decreased and throughout the greater part of the proliferative compartment was between 0.40 and 0.50 hr. At the top of the proliferative compartment an increase in t _m was demonstrated.
If the value of 0.43 hr for the whole crypt column is correct, then one argument for postulating the formation of metabolic DNA during differentiation in the small bowel epithelium of the rat becomes invalid. Variations in t _m within the crypt have been shown to increase the values of cell velocity obtained from cumulative birth rate diagrams. Finally further evidence has been presented for the existence of a slowly dividing subpopulation of cells at the base of the crypt. These cells may be important in crypt repopulation after damage with phase specific anti-tumour drugs.     

Metabolism of Imidazole by a Pseudomonad

Intermediates formed during the microbial degradation of imidazole, namely 4(5)-imidazolone, formiminoglycine, and possibly glycine, are similar to those formed during metabolism of imidazole derivatives.     

The relationship of root-cap slimes to proteins

1. The patterns of incorporation of radioactivity from d-[U-(14)C]glucose into the pectic components of sections of sycamore roots changed so that sections nearer the tip incorporated relatively more label into arabinose and galactose compared with uronic acid. 2. Radioactive maize root-cap slime was prepared and found to contain three water-soluble component polymers which were electrophoretically (i) neutral, (ii) weakly acidic and (iii) strongly acidic at pH6.5. The neutral component was a glucan. The other components, which could be degraded by trans-elimination, consisted of an acidic backbone chain composed of galacturonic acid and glucose, attached to which were different proportions of neutral sugars. Arabinose, galactose and fucose, the main neutral sugars of the weakly and strongly acidic materials, were absent from the neutral fraction. 3. Fucose was a major sugar in maize-root slime and in a slime of similar composition synthesized by a maize callus of shoot origin. Only trace amounts were found in sycamore, pea and wheat root tips, and in pectin prepared from maize roots and coleoptiles. A high proportion of fucose is therefore a chemical characteristic of maize slime, and slime synthesis indicated a state of differentiation of the tissue. 4. The similarity between the slime and pectin is discussed; slime is a form of pectin modified in such a way as to provide a hydrated protective coating around the root tip.     

Temperature-sensitive Mutants of Respiratory Syncytial Virus

Four conditional-lethal temperature-sensitive mutants of RS virus were detected among the progeny of 454 plaques derived from virus grown in the presence of 10(-4)m 5-fluorouridine. These mutants were stable (reversion frequency, 10(-5.0) or less and failed to produce plaques at 38 or 39 C. Plaquing efficiency was depressed 100-fold or more at 37 C. Variable suppression of growth at the restrictive temperature of 39 C was observed, ranging from 16-fold to complete suppression. The temperature-sensitive defect of three of the mutants appeared to affect functions which were expressed late in the replicative cycle. One of the mutants produced atypical nonsyncytial plaques.     

The fine structure of the sense organs of the cephalopod mollusc Nautilus

Summary The structure of the rhinophore, digital tentacles, post-ocular tentacles and the eye of Nautilus macromphalus are described. The rhinophore is composed of mucous cells, ciliated cells, and flask-shaped ciliated cells. The latter are probably olfactory receptors. The digital tentacles are composed of mucous cells and pigmented cells. Motor-end-plates found in the muscle layer below the epithelium of the digital tentacles are similar to those described in other cephalopods. The post-ocular tentacle contains receptor cells that bear macrocilia. These may be mechanoreceptors. The retina is composed of retinula cells and supporting cells. A complex rhabdom is formed at the distal ends of the retinula cells. The supporting cells send processes up between these rhabdoms. Both types of cells contain pigment granules but the retinula cell has a complex membranous structure in its perikaryon. No synapses were found at the bases of the retinula cells. At the side of the retina are mucous cells that are presumed to produce the jelly-like substance that fills the inside of the eye in life. The likely function of the eye is discussed and it is suggested that it is capable of simple discriminations. It is suggested that the sense organs are probably comparatively unchanged from those of fossil nautiloids. Acknowledgements. This paper is dedicated to the late Dr. Yves Merlet who collected the nautiluses used in this study.We would like to thank Prof. J. Z. Young for all his support and encouragement. The Royal Society, The Percy Sladen Memorial Fund, and University College, London, provided the financial support that enabled one of us (V.C.B.) to collect nautiluses. The Science Research Council, U.K., provided the electron microscope used in the major part of the study and a grant to one of us (V.C.B.). We would also like to thank Prof. J. B. Gilpin-Brown who provided Fig. 1, Dr. R. Catala, for aquarium facilities, Mr. M. P. Legand and the Institut Français d'Oceanie, Noumea, New Caledonia, for laboratory facilities, Dr. J.-M. Bassot and Dr. Anna Bidder for advice on catching and preserving nautiluses, Mrs. Judy Parkes and Mr. M. Barker for photographic assistance, and Miss J. Date for secretarial assistance.