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Markus Fischer

Publications and source records attributed to Markus Fischer.

78 records · Page 5Linked to original sources
PubMed2002

Studies on the reaction mechanism of riboflavin synthase: X-ray crystal structure of a complex with 6-carboxyethyl-7-oxo-8-ribityllumazine.

Riboflavin synthase catalyzes the disproportionation of 6,7-dimethyl-8-ribityllumazine affording riboflavin and 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione. We have determined the structure of riboflavin synthase from Schizosaccharomyces pombe in complex with the substrate analog, 6-carboxyethyl-7-oxo-8-ribityllumazine at 2.1 A resolution. In contrast to the homotrimeric solution state of native riboflavin synthase, we found the enzyme to be monomeric in the crystal structure. Structural comparison of the riboflavin synthases of S. pombe and Escherichia coli suggests oligomer contact sites and delineates the catalytic site for dimerization of the substrate and subsequent fragmentation of the pentacyclic intermediate. The pentacyclic substrate dimer was modeled into the proposed active site, and its stereochemical features were determined. The model suggests that the substrate molecule at the C-terminal domain donates a four-carbon unit to the substrate molecule bound at the N-terminal domain of an adjacent subunit in the oligomer.

Amino Acid Sequence↗
PubMed2002

Biosynthesis of riboflavin: 6,7-dimethyl-8-ribityllumazine synthase of Schizosaccharomyces pombe.

A cDNA sequence from Schizosaccharomyces pombe with similarity to 6,7-dimethyl-8-ribityllumazine synthase was expressed in a recombinant Escherichia coli strain. The recombinant protein is a homopentamer of 17-kDa subunits with an apparent molecular mass of 87 kDa as determined by sedimentation equilibrium centrifugation (it sediments at an apparent velocity of 5.0 S at 20 degrees C). The protein has been crystallized in space group C2221. The crystals diffract to a resolution of 2.4 A. The enzyme catalyses the formation of 6,7-dimethyl-8-ribityllumazine from 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione and 3,4-dihydroxy- 2-butanone 4-phosphate. Steady-state kinetic analysis afforded a vmax value of 13 000 nmol.mg-1.h-1 and Km values of 5 and 67 microm for 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione and 3,4-dihydroxy-2-butanone 4-phosphate, respectively. The enzyme binds riboflavin with a Kd of 1.2 microm. The fluorescence quantum yield of enzyme-bound riboflavin is < 2% as compared with that of free riboflavin. The protein/riboflavin complex displays an optical transition centered around 530 nm as shown by absorbance and CD spectrometry which may indicate a charge transfer complex. Replacement of tryptophan 27 by tyrosine or phenylalanine had only minor effects on the kinetic properties, but complexes of the mutant proteins did not show the anomalous long wavelength absorbance of the wild-type protein. The replacement of tryptophan 27 by aliphatic amino acids substantially reduced the affinity of the enzyme for riboflavin and for the substrate, 5-amino-6-ribitylamino-2,4(1H,3H)-pyrimidinedione.

Amino Acid Sequence↗
PubMed2002

Localization of two enzymes of the tetrahydrobiopterin biosynthetic pathway in embryonic chick retina.

Tetrahydrobiopterin (BH4) is an essential co-factor for the biosynthesis of catecholamine-type neurotransmitters and of nitric oxide (NO). The expression of the enzymes catalyzing the first two steps of the BH4 biosynthetic pathway was studied in the developing chicken retina by in situ hybridization and immunocytochemistry. GTP-cyclohydrolase-I (GTP-CH-I) and 6-pyruvoyl-tetrahydropterin synthase (PTPS) were already expressed in the undifferentiated and proliferating retina of E7. At stage E11 both enzymes were expressed in photoreceptors, amacrine cells, displaced amacrine cells, and ganglion cells, and in the plexiform layers in which synaptic connections take place. At stage E18 the labeling was comparable to E11 but appeared to be more concentrated in photoreceptors and ganglion cells.

Animals↗
PubMed2002

Experimental life-history evolution: selection on the allocation to sexual reproduction and its plasticity in a clonal plant.

Allocation to sexual reproduction is an important life-history trait in clonal plants. Different selection pressures between competitive and competition-free environments are likely to result in the evolution of specialized genotypes and to maintain genetic variation in reproductive allocation. Moreover, selection may also result in the evolution of plastic allocation strategies. The necessary prerequisite for evolution, heritable genetic variation, can best be studied with selection experiments. Starting from a base population of 102 replicated genotypes of the clonal herb Ranunculus reptans, we imposed selection on the proportion of flowering rosettes in the absence of competition (base population: mean = 0.391, broad-sense heritability = 0.307). We also selected on the plasticity in this trait in response to competition with a naturally coexisting grass in a parallel experiment (base population: 14% lower mean in the presence of competition, broad-sense heritability = 0.072). After two generations of bidirectional selection, the proportion of flowering rosettes was 26% higher in the high line than in the low line (realized heritability +/- SE = 0.205 +/- 0.017). Moreover, genotypes of the high line had 11% fewer carpels per flower, a 22% lower proportion of rooted rosettes, and a 39% smaller average distance between rosettes within a clone. In the second experiment, we found no significant responses to selection for high and low plasticity in the proportion of flowering rosettes (realized heritability +/- SE = -0.002 +/- 0.013). Our study indicates a high heritability and potential for further evolution of the proportion of flowering rosettes in R. reptans, but not for its plasticity, which may have been fixed by past evolution at its current level. Moreover, our results demonstrate strong genetic correlations between allocation to sexual reproduction and other clonal life-history characteristics.

Biological Evolution↗
PubMedDate not supplied

Dual modality of 18F-fluorodeoxyglucose-positron emission tomography/computed tomography in patients with cervical carcinoma of unknown primary.

OBJECTIVE: To evaluate an optimized F-18-flurodeoxyglucose-positron emission tomography/computed tomography (FDG-PET/CT) acquisition protocol for head and neck cancer and assess the usefulness of combined FDG-PET/CT in locating unknown primary tumors in patients with biopsy-proven cervical lymph node metastases. SUBJECTS AND METHODS: Twenty-one patients with cervical lymph node metastases of unknown primary tumors underwent staging with FDG-PET/CT. The images of FDG-PET alone, CT alone, FDG-PET/CT read side by side and fused and FDG-PET/CT were evaluated separately by 2 physicians. Imaging results were correlated with either histology (n = 14) or clinical follow-up (n = 7). RESULTS: On the fused FDG-PET/CT images, primary tumors were identified in 12 patients (57%); with FDG-PET alone and FDG-PET and CT read side by side 11 (52%) primary tumors were found while CT alone identified 5 (23%) primary tumors. CONCLUSION: Our data indicate that fused FDG-PET and CT images increased the sensitivity of detecting carcinoma of unknown primary (CUP) tumors compared to CT alone, but not to FDG-PET alone or FDG-PET and CT read side by side. Hence accurate fusion of functional and morphologic data by FDG-PET/CT is a promising imaging modality in the clinical workup of patients with cervical CUP tumors.

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