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Ohne Zusammenfassung     

A more accurate method for measurement of tuberculocidal activity of disinfectants.

The current Association of Official Analytical Chemists method for testing tuberculocidal activity of disinfectants has been shown to be inaccurate and to have a high degree of variability. An alternate test method is proposed which is more accurate, more precise, and quantitative. A suspension of Mycobacterium bovis BCG was exposed to a variety of disinfectant chemicals and a kill curve was constructed from quantitative data. Data are presented that show the discrepancy between current claims, determined by the Association of Official Analytical Chemists method, of selected commercially available products and claims generated by the proposed method. The effects of different recovery media were examined. The data indicated that Mycobacteria 7H11 and Middlebrook 7H10 agars were equal in recovery of the different chemically treated cells, with Lowenstein-Jensen agar having approximately the same recovery rate but requiring incubation for up to 3 weeks longer for countability. The kill curves generated for several different chemicals were reproducible, as indicated by the standard deviations of the slopes and intercepts of the linear regression curves.     

Effect of amino acid substitutions and deletions on the thermal stability, the pH stability and unfolding by urea of bovine calbindin D9k

The influence of amino acid substitutions and deletions on the stability of bovine calbindin D9k, the smallest protein known with a pair of EF-hand calcium-binding sites, has been studied using circular dichroism and ultraviolet absorption spectroscopy. The five modifications are confined to one of the two Ca2+ -binding sites. The Ca2+-loaded forms of the wild-type and mutant calbindins are too stable to be significantly denatured by heating at 90 degrees C or by adding 8 M urea. For the Ca2+-free (apo) forms thermal unfolding appears to be only half complete at 90 degrees C, while denaturation is complete in 7-8 M urea. Four of the mutant proteins show reduced resistance towards unfolding by urea, but one of the modified proteins (Glu-17----Gln) shows an increased stability, presumably because of a reduced electrostatic repulsion in the native state. According to X-ray crystallographic data the OH group of the single tyrosine of calbindin (Tyr-13) is hydrogen-bonded to the carboxyl group of Glu-35, thus linking the two alpha helices flanking the N-terminal Ca2+ site. The pK of ionization of the Tyr-13 hydroxyl group was over 13 for calcium forms of the wild-type protein, between 12.3 and 12.8 for the calcium form of three mutants and between 11.5 and 11.7 for the apoproteins. Significant differences in pH stability between wild type and mutants were observed in the calcium forms, but were not apparent in the apo forms.     

Saturation transfer EPR spectroscopy on spin-labeled muscle fibers using a loop-gap resonator.

Previously, saturation transfer (ST-EPR) studies of biomolecular dynamics have involved the use of a resonant cavity and the V'2 display (absorption, second harmonic, out of phase). In the present study, we replaced the resonant cavity with a loop-gap resonator and used the U'1 display (dispersion, first harmonic, out of phase) to study spin-labeled muscle fibers. The new resonator and display showed several advantages over those previously used. It produced virtually noiseless U'1 spectra on a 0.4 microliter sample using a 4 min scan; previous U'1 experiments on spin-labeled muscle, using a conventional rectangular cavity, resulted in an unacceptably low signal-to-noise ratio. The high filling factor of the resonator facilitated the study of these extremely small fiber bundles and permitted high microwave field intensities to be achieved at much lower incident microwave power levels, thus greatly enhancing the signal-to-noise ratio in U'1 experiments. This reduction in the noise level made it possible to benefit from the other advantages of U'1 over V'2, such as stronger signals, simpler line shapes, and simpler data analysis. For these muscle fiber samples, the resulting sensitivity (signal/noise/sample volume) of the U'1 signals was greater than 100 times that of V'2 signals obtained in a conventional cavity. Another advantage of the U'1 display is that signals from weakly immobilized probes, i.e., probes that have nanosecond rotational mobility relative to the labeled protein (myosin), are greatly suppressed relative to strongly immobilized probes. This reduces the ambiguity of spectral analysis, and eliminates the need for chemical treatments [e.g., using K3Fe(CN)6] that were previously required in muscle fibers and other systems. Further suppression of this weakly immobilized component was achieved in U'1 spectra by increasing the microwave power and decreasing the field modulation frequency.     

Genetisch kontrollierte Varianten der NADH-Diaphorase

Zusammenfassung Die NADH-Diaphorase wurde an 725 gesunden Probanden mit Hilfe der Stärkegelelektrophorese untersucht. Zwei verschiedene Varianten wurden beobachtet: eine heterozygot schnelle (DIA 2-1) und eine heterozygot langsame (DIA 3-1). Die Genhäufigkeiten sind: DIA²=0,0021; DIA³=0,0007.

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Genetically determined variants of NADH-diaphorase

Summary By means of starchgel-electrophoresis a screening for variants of NADH-Diaphorase was carried out within a sample of 725 healthy probands. Two kinds of genetically determined variants have been observed: a heterozygous phenotype with greater mobility (DIA 2-1) and a heterozygous phenotype with slower mobility (DIA 3-1). The gene-frequencies are estimated so far as 0.0021 (DIA²) and 0.0007 (DIA³).

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Mit Unterstützung durch die Deutsche Forschungsgemeinschaft.     

Beitrag zur Populationsgenetik der Pyruvatkinase menschlicher Erythrocyten

Summary In 214 healthy young Germans the activity of Pyruvatekinase from red blood cells has been determined. Three persons had values in the heterozygote range between 10.0 and 20.0 U. Suggesting a 2-allele-model the frequency of the three phenotypes in the German population can be calculated as followed: PK(A)=98.6%, PK(AB)=1.4%, PK(B)=0.005%.No correlation could be found between the distribution of blood-and serum-groups and the enzyme-activity.

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Direktor: Prof. Dr. med. G. W. Löhr

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Direktor: Prof. Dr. med. G. G. Wendt

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Mit dankenswerter Unterstützung durch die Deutsche Volkswagen-Stiftung.

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Herrn Professor Dr.H. E. Bock, Tübingen, zum 65. Geburtstag gewidmet. Wesentliche Teile dieser Arbeit werden von Olaf Praetsch der Medizinischen Fakultät als Dissertation vorgelegt.     

mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Crystallization and preliminary X-ray crystallographic analysis of squalene-hopene cyclase from Alicyclobacillus acidocaldarius.

The membrane-associated protein squalene-hopene cyclase from Alicyclobacillus acidocaldarius was overexposed in Escherichia coli and purified by ion exchange and gel permeation chromatography. Crystals of three interrelated forms were grown by vapor diffusion under identical conditions. The crystals diffract to about 2.3 A resolution, but they are unstable in the X-ray beam. An interpretable heavy-atom derivative was obtained.     

De novo trisomy 4pter→q21

Summary Clinical and cytogenetic findings in an infant girl with multiple congenital anomalies, principally anophthalmia, are presented. The patient's karyotype was 47,XX, +del(4)(pterq21), the largest partial trisomy of chromosome 4 reported. The possible mechanism of the origin of this abnormality is discussed.To whom offprint requests should be sent