KARYOKINESIS OF SOMATIC NUCLEI OF<i>NEUROSPORA CRASSA</i>: III. THE JUVENILE AND MATURATION CYCLES (FEULGEN AND CRYSTAL VIOLET STAINING)

Weijer, J.; Koopmans, A.; Weijer, D.L.

doi:10.1139/g65-020

Cited by 28 publications

(12 citation statements)

References 27 publications

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“…2. The amino acid sequence agrees with the chemically determined sequence [39]. The initiating formylmethionine is not removed post-translationally.…”

Section: Dna-sequence Analysissupporting

confidence: 77%

Substitution of Arg214 at the substrate‐binding site of p, ‐hydroxybenzoate hydroxylase from Pseudomonas fluorescens

Berkel

Westphal

Eschrich

et al. 1992

European Journal of Biochemistry

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The gene encoding p-hydroxybenzoate hydroxylase from Pseudomonas,fluorescens was cloned in Escherichia coli to provide DNA for mutagenesis studies on thc protein product. A plasmid containing a 1.65-kbp insert of P. fluorescens chromosomal DNA was obtained and its nucleotide sequence determined. The DNA-derived amino acid sequence agrees completely with the chemically determined amino acid sequence of the isolated protein. The enzyme is strongly expressed under influence of the vector-encoded lac promotor and is purified to homogeneity in a simple three-step procedure.The relation between substrate binding, the effector role of substrate and hydroxylation efficiency was studied by use of site-directed rnutagenesis. Arg214, in ion-pair interaction with the carboxy moiety of p-hydroxybenzoate, was replaced with Lys, Gln and Ala, respectively. The affinity of the free enzymes for NADPH is unchanged, whereas the affinity for the aromatic substrate is strongly decreased. For enzymes Arg214jAla and Arg214+Gln, the effector role of substrate is lost. For enzyme Arg214+Lys, binding of p-hydroxybenzoate highly stimulates the rate of flavin reduction.In the presence of substrate or substrate analogues, the reduced enzyme Arg214jLys fails to stabilize the 4a-hydroperoxyflavin intermediate, essential for efficient hydroxylation. Like the wild-type, enzyme Arg214+Lys is susceptible to substrate inhibition. From spectral and kinetic results it is suggested that secondary binding of the substrate occurs at the re side of the flavin, where the nicotinamide moiety of NADPH is supposed to bind.p-Hydroxybenzoate hydroxylase is a member of the class of flavin-dependent monooxygenases [I]. The enzyme catalyzes the conversion of 4-hydroxybenzoate to 3,4-dihydroxybenzoate, an intermediate step in the degradation of aromatic compounds in soil bacteria [2]. Like the majority of the flavoprotein aromatic hydroxylases, p-hydroxybenzoate hydroxylase shows a narrow substrate specificity [l, 31. The substrate also acts as an effector, strongly stimulating the reduction of protein-bound FAD by NADPH [3,4].p-H ydroxybenzoate hydroxylase from Pseudomonas Juorescens is one of the best studied flavoproteins [l]. Catalytic and biophysical properties have been investigated in detail [5 -81 and high-resolution crystal structures of the enzyme in complex with substrate or product are available [9 -121. The structure of the free enzyme is less well resolved [13], and the binding mode of NADPH remains to be elucidated [14].

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“…2. The amino acid sequence agrees with the chemically determined sequence [39]. The initiating formylmethionine is not removed post-translationally.…”

Section: Dna-sequence Analysissupporting

confidence: 77%

Substitution of Arg214 at the substrate‐binding site of p, ‐hydroxybenzoate hydroxylase from Pseudomonas fluorescens

Berkel

Westphal

Eschrich

et al. 1992

European Journal of Biochemistry

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“…1c) and the young terminal vesicles are charged with many well-defined nuclei (Benjamin, 1959, Plate la). Such elongate nuclei are reminiscent of the filamentous nuclei reported in Neurospora and Gelasinospora by Dowding and Weijer (1962) and Weijer, Koopmans, and Weijer (1965). la).…”

supporting

confidence: 66%

The Merosporangium

Benjamin

1966

Mycologia

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“…But information as to how disjunction is achieved is not given. In Ascomycetes an elongated nucleus dividing by longitudinal fission is described by some authors: in Neurospora crassa by Weijer et al (1963by Weijer et al ( , 1965 and by Namboodiri & Lowry (1967, 1968, in Nannizia incurvata by Hejtm~inkov~i-Uhrov~i & Hejtmanek (1968), in Trichophyton Vanbreuseghemii by Hejtm~inek & Hejtm~inkov~i-Uhrova (1968), and in Penicillium species by Laane (1967Laane ( , 1969.…”

Section: Discussionmentioning

confidence: 98%

“…Harper called it 'central body', Colson 'lateral granule'. Several other investigators noticed this lateral granule, both in ascus nuclei and in resting hyphal nuclei (McClintock, 1945;Singleton, 1953;Olive, 1953;Ward & Ciurysek, 1962;Schopfer et al, 1963;Weijer et al, 1963Weijer et al, , 1965Berlin & Bowen, 1964;Wilson & Aist, 1967;Robinow & Caten, 1969;Aist, 1969). In electron micrographs of fungal Ascomycetes two 'centriolar plaques' are also described (Wells, 1970;Zickler, 1970;Van Winkel et al, 1971), sometimes connected by an intranuclear spindle.…”

Section: Centriolesmentioning

confidence: 93%

A cytological study of Nematospora coryli pegl

Koopmans

1977

Genetica

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KARYOKINESIS OF SOMATIC NUCLEI OFNEUROSPORA CRASSA: III. THE JUVENILE AND MATURATION CYCLES (FEULGEN AND CRYSTAL VIOLET STAINING)

Cited by 28 publications

References 27 publications

Substitution of Arg214 at the substrate‐binding site of p, ‐hydroxybenzoate hydroxylase from Pseudomonas fluorescens

Substitution of Arg214 at the substrate‐binding site of p, ‐hydroxybenzoate hydroxylase from Pseudomonas fluorescens

The Merosporangium

A cytological study of Nematospora coryli pegl

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