Evolution of structure and substrate specificity ind-alanine:d-Alanine ligases and related enzymes

Thed-alanine:d-alanine-ligase-related enzymes can have three preferential substrate specificities. Usually, these enzymes synthesized-alanyl-d-alanine. In vancomycin-resistant Gram-positive bacteria, structurally related enzymes synthesized-alanyl-d-lactate or Dalanyl-d-serine. The sequence of internal fragments of eight structurald-alanine:d-alanine ligase genes from enterococci has been determined. Alignment of the deduced amino acid sequences with those of other related enzymes from Gram-negative and Gram-positive bacteria revealed the presence of four distinct sequence patterns in the putative substrate-binding sites, each correlating with specificity to a particular substrate (d-alanine:d-lactate ligases exhibited two patterns). Phylogenetic analysis showed different clusters. The enterococcal subtree was largely superimposable on that derived from 16S rRNA sequences. In lactic acid bacteria, structural divergence due to differences in substrate specificity was observed. Glycopeptide resistance proteins VanA and VanB, the VanC-type ligases, and Dd1A and DdlB from enteric bacteria andHaemophilus influenzae constituted separate clusters. Correspondence to: P. Courvalin     

Oxygen starvation induces cell death in Candida shehatae fermentations of d-xylose, but not d-glucose

Candida shehatae cells, cultivated on d-glucose and d-xylose, were subjected to a shift from fully aerobic to anaerobic fermentative conditions. After anaerobic conditions were imposed, growth was limited to approximately one doubling or less as C. shehatae rapidly entered a stationary phase of growth. Following the shift to anoxia, cell viability rapidly declined and the total cell volume declined in the d-xylose fermentations. Moreover, the cell volume distribution shifted to smaller volumes. Cell viability, measured by plate counts, declined nine times faster for d-xylose fermentations than for d-glucose fermentations. Anaerobic growth did not occur on either d-glucose or d-xylose. Selected vitamins and amino acids did not stimulate anaerobic growth in C. shehatae, but did enhance anaerobic growth on d-glucose in S. cerevisiae. The decline in cell viability and lack of anaerobic growth by C. shehatae were attributed to oxygen deficiency and not to ethanol inhibition. The results shed light on why C. shehatae anaerobic fermentations are not currently practical and suggest that research directed towards a biochemical understanding of why C. shehatae can not grow anaerobically will yield significant improvements in ethanol fermentations from d-xylose. Received 26 October 1998 / Received revision: 26 January 1999 / Accepted: 12 February 1999     

A taxonomical revision ofThymus sect.Teucrioides (Lamiaceae)

TheThymus teucrioides Boiss. & Spruner aggregate is revised and the following new taxa, all from the alpine zone in the Greek mountains, are described:Th. leucospermus Hartvig from the calcareous mountains of Pindhos and Mt Parnassos in Sterea Ellas,Th. rechingeri Hartvig with the subsp.macrocalyx Hartvig from calcareous mountains in Sterea Ellas and N Peloponnissos, andTh. teucrioides subsp.alpinus Hartvig from the serpentine areas of N Pindhos. In the variableTh. teucrioides s. str. many characters have turned out to be markedly geographically correlated and many local populations can be distinguished by a particular combination of characters.Dedicated to Prof.K. H. Rechinger on the occasion of his 80th birthday.     

Structure of the peptidoglycans of Moraxella glucidolytica and Moraxella lwoffi grown on hydrocarbons

The peptidoglycans of Moraxella glucidolytica and Moraxella lwoffi grown on aliphatic hydrocarbons were isolated. They contained muramic acid. glucosamine, alanine, d-glutamic acid and meso-diaminopimelic acid in a molar ratio of about 0.5:0.5:1.6:1.0:1.0 (M. glucidolytica) and 0.8:0.7:1.3:1.0:1.0 (M. lwoffi).The peptidoglycans were lysozyme-resistant. However, when treated with formamide, they could be partially degraded by lysozyme. The fragments were purified and their structure determined. In both strains, the peptide subunits consisted mainly of tripeptides (l-Ala-d-Glu-meso-DAP) and tetrapeptides (l-Ala-d-Glu-meso-DAP-d-Ala), most of them being directly cross-linked. It is concluded that in both strains the primary structures of the peptidoglycans are closely related.     

Peptidoglycan types and cytochrome patterns of strains of Oerskovia turbata and O. xanthineolytica

Determination of the primary structure of the peptidoglycan of 15 strains of Oerskovia showed that three different peptidoglycan types occur. Oerskovia xanthineolytica strains contain the l-Lys-d-Ser--d-Asp type, whereas Oerskovia turbata strains show the new peptidoglycan types l-Lys-l-Thr--d-Asp or l-Lys-l-Thr--d-Glu, respectively. Research on the cytochromes of Oerskovia revealed the presence of a, b and c types. O. turbata can be clearly distinguished from O. xanthineolytica by the occurrence of cytochrome a ₁ in cells, isolated from the stationary phase. The following conclusions were made: O. turbata and O. xanthineolytica can be clearly separated on the basis of different peptidoglycan types and cytochrome patterns. This distinction is in perfect correlation with the classical separation method of O. turbata and O. xanthineolytica on the basis of xanthine degradation. l-Lys-d-Ser--d-Asp peptidoglycan type does not only occur in O. xanthineolytica but also in some coryneform bacteria such as Corynebacterium manihot (Fiedler et al. 1970), Cellulomonas cartae (Stackebrandt et al. 1978; Stackebrandt and Kandler 1980), Brevibacterium fermentans and Nocardia cellulans.This paper is respectively dedicated to Professor Dr. O. Kandler, on the occasion of his 60th birthday     

Dois novos taxa de Sphaeropsidaceae sôbre Palmaceae

Summary This paper deals with two new fungi studied on Palmaceae:Asteromella cocoes Batista &Bezerra n. sp. on leaves ofCocos nucifera L. andAsteropsis insectorum Batista &Bezerra n. sp. onCoccideae scales on leaves ofOenocarpus distichus Mart.

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Zusammenfassung Diese Arbeit beschreibt zwei neue Pilze, welche auf Palmaceae studiert wurden:Asteromella cocoes Batista &Bezerra n. sp. auf Blättern vonCocos nucifera L. undAsteropsis insectorum Batista &Bezerra n. sp. aufCoccideae Schuppen auf Blättern vonOenocarpus distichus Mart.

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Dedicado à Memória do Prof.Raffaele Ciferri     

New and noteworthyHabenaria spp. (Orchidaceae) from New Guinea and adjacent islands

5 new taxa ofHabenaria, namelyH. bougainvillae, H. elongata R. Br. var.leptophylla, H. ensigera, H. rechingeri andH. trichoglossa, are described and illustrated, with reference to affinities to related Australian and Indo-malayan species. The occurrence in New Guinea of severalHabenaria spp. typical for a savanna-like vegetation, led to look more thoroughly at these taxa:H. elongata R. Br. andH. ochroleuca R. Br., considered so far to be endemic in Northern Australia, andH. khasiana Hook. f., hitherto only known from southeastern Asia.Studies in the subtribeHabenariinae Bentham (Orchidaceae), 2.—Part 1: Candollea34, 357 (1979).Dedicated to Hofrat Prof. DrKarl Heinz Rechinger on the occasion of his 80th birthday. — On April 30, in the year 1927, the author and his brotherOtto Renz metKarl Heinz Rechinger on a small steam-boat in a stormy Aegaen Sea, travelling from Piraeus to the Northern Sporades Islands:Karl Heinz with destination to Chelidromia, the author andO. Renz to Skopelos. By this lucky chance a lasting friendship began.     

Overproduction of d-hydantoinase and carbamoylase in a soluble form in Escherichia coli

The production of d-hydantoinase and carbamoylase from Agrobacterium radiobacter NRRL B11291 using T7 and trc promoters, respectively, was found to cause protein aggregates in Escherichia coli. We initiated a systematic study aimed at overproducting these two proteins in a soluble form. As a result, the protein aggregate from carbamoylase overproduction could be alleviated with the aid of GroEL/GroES. In contrast, the production of a high level of d-hydantoinase in an active form can be achieved at low temperature (25 °C) or by the coproduction of DnaJ/DnaK. Overall, with such approaches both recombinant proteins gain more than a four-fold increase in enzyme activity. In addition, by fusion with thioredoxin, d-hydantoinase activity can be increased 25% more than the unfused counterpart in the presence of DnaJ/DnaK. These results indicate the success of our approaches to overproducing d-hydantoinase and carbamoylase in a soluble form in E. coli. Received: 26 November 1999 / Received revision: 28 February 2000 / Accepted: 10 March 2000     

Larval and pupal parasites ofLymantria dispar and notes on parasites of otherLymantriidae [Lep.] in Poland 1975

Larvae and pupae ofLymantria dispar (L.) were collected periodically from infestations on Linden trees at Nieborów near Skierniewice and at Alexandrowa, south of Poznan, Poland during May–July 1975. The following parasites were recovered:Braconidae: Apanteles liparidis (Bouché),A. melanoscelus (Ratzeburg),A. porthetriae Muesebeck,Meteorus pulchricornis (Wesmael);Ichneumonidae: Coccygomimus instigator (F.),Hyposoter tricoloripes (Viereck),Phobocampe n. sp.,P. disparis (Viereck);Tachinidae: Blepharipa pratensis (Meigen),Blondelia nigripes (Fallén),Carcelia separata (Rondani),Compsilura concinnata (Meigen),Parasetigena silvestris (Robineau-Desvoidy),Palexorista sp. Similar but less extensive collections were made of larvae ofOrgyia antiqua (L.). These yieldedApanteles sp.,A. melanoscelus, Tramosema rostrale (Brischke),Meloborus sp.,B. nigripes andP. silvestris; larvae ofLeucoma salicis (L.) yieldedA. melanoscelus andMeteorus versicolor (Wesmael). OnlyBlondelia sp. was recovered from the few larvae ofPorthesia similis (Fuessly) collected.

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Résumé Des chenilles et des chrysalides deLymantria dispar (L.) ont été ramassées périodiquement en Pologne de mai à juillet 1975 dans les peuplements de tilleuls infestés, à Nieboróv près de Skierniewice et à Alexandrowa, au sud de Poznan. Les parasites obtenus ont été les suivants: lesBraconidae: Apanteles liparidis (Bouché),A. melanoscelus (Ratzeburg),A. porthetriae Muesebeck,Meteorus pulchricornis (Wesmael); lesIchneumonidae: Coccygomimus instigator (F.),Hyposoter tricoloripes (Viereck),Phobocampe n. sp.,P. disparis (Viereck); lesTachinidae: Blepharipa pratensis (Meigen),Blondelia nigripes (Fallén),Carcelia separata (Rondani),Compsilura concinnata (Meigen),Parasetigena silvestris (Robineau-Desvoidy), etPalexorista sp. Des collectes du même type mais moins importantes de chenilles d’Orgyia antiqua (L.) ont permis d’obtenir les parasites suivants:Apanteles sp.,A. melanoscelus. Tranosema rostrale (Brischke),Meloborus sp.,B. nigripes, etP. silvestris. Des chenilles deLeucoma salicis (L.) ont donnéA. melanoscelus etMeteorus versicolor (Wesmael). SeulBlondelia sp. a été obtenu à partir de quelques chenilles dePorthesia similis (Fuessly).

     

Substrate Selectivity and pH Dependence of KAAT1 Expressed in Xenopus laevis Oocytes

When expressed in Xenopus oocytes KAAT1 increases tenfold the transport of l-leucine. Substitution of NaCl with 100 mm LiCl, RbCl or KCl allows a reduced but significant activation of l-leucine uptakes. Chloride-dependence is not strict since other pseudohalide anions such as thyocyanate are accepted. KAAT1 is highly sensitive to pH. It can transport l-leucine at pH 5.5 and 8, but the maximum uptake has been observed at pH 10, near to the physiological pH value, when amino and carboxylic groups are both deprotonated. The pH value mainly influences the V _max in Na⁺ activation curves and l-leucine kinetics. The kinetic parameters are K _mNa = 4.6 ± 2 mm, V _maxNa = 14.8 ± 1.7 pmol/oocyte/5 min for pH 8.0 and K _mNa = 2.8 ± 0.7 mm, V _maxNa = 31.3 ± 1.9 pmol/oocyte/5 min for pH 10.0. The kinetic parameters of l-leucine uptake are: K _m = 120.4 ± 24.2 μm, V _max = 23.2 ± 1.4 pmol/oocyte/5 min at pH 8.0 and K _m = 81.3 ± 24.2 μm, V _max = 65.6 ± 3.9 pmol/oocyte/5 min at pH 10.0. On the basis of inhibition experiments, the structural features required for KAAT1 substrates are: (i) a carboxylic group, (ii) an unsubstituted α-amino group, (iii) the side chain is unnecessary, if present it should be uncharged regardless of length and ramification. Received: 27 April 1999/Revised: 10 January 2000     

Delimitation and classification ofBromus fasciculatus (Poaceae)

The syntypes ofBromus madritensis var.delilei Boiss. comprise two different elements: specimens ofB. haussknechtii Boiss. and ofB. fasciculatus C. Presl s.l. By its lectotypification and on the basis of new morphological characters, the validation ofB. fasciculatus subsp.delilei (Boiss.)H. Scholz 1971 was substantiated. This subspecies represents the eastern marginal segregate of the south-Mediterranean species.B. fasciculatus var.alexandrinus Thell. is a minor variant of the typical subspecies.Dedicated to Prof.K. H. Rechinger on the occasion of his 80th birthday.     

Production of d-amino acids using immobilized d-hydantoinase from lentil, Lens esculenta, seeds

d-Hydantoinase from the lentil, Lens esculenta, seed is quite unstable, and has been immobilized on Diethyl amino ethyl (DEAE) cellulose by an adsorption and cross-linking method. The immboilized d-hydantoinase exhibited 80% enzyme activity and contained 86% protein. The immobilization of the enzyme preparation does not change its optimum pH, temperature or affinity constant, but increases its shelf-life, thermostability and stability in various organic solvents. This immobilized d-hydantoinase can be used effectively for the production of d-amino acids from the corresponding hydantoins and may therefore be of use in the chemical and pharmaceutical industries. Received: 28 April 1998 / Received last revision: 10 July 1998 / Accepted: 10 July 1998     

Contributo alla sistematica degli attinomiceti: XVII actinomices scabies (Thaxter) Waksman

Riassunto Gli autori studiano un ceppo di attinomicete che essi identificano comeA. scabies. Con l'occasione essi precisano le caratteristiche diagnostiche di questa specie secondo quanto risulta da un esame critico della bibliografia in argomento, mettendo in evidenza che i ceppi collezionati sotto questo nome nelle varie micoteche hanno spesso caratteristiche differenti. La base di questo equivoco è, nelle considerazioni degli autori, da trovare nel riferimento patogenetico, cioè, in altri termini, nel voler considerare la malattia dei tuberi scabbiosi come provocata da un'unica specie.Dopo aver dato le caratteristiche delle specie presentano il seguente quadro tassonomico: Actinomyces scabies (Thaxter 1890)Gussov (1914) emendamentoWaksman (1919)=Oospora scabies Thaxter (1890)Diagnosi: vedi pag. 273.Habitat: frequente nel Nord America; isolato per la prima volta in Europa daSpalla eComaschi da tuberi scabbiosi provenienti dalla Germania. Actinomyces scabies varietàanglica Baldacci eBoncompagni =Actinomyces scabies Thaxter emendamentoMillard eBurr.Differisce dalla specie per il micelio vegetativo grigio, il micelio aereo spiccatamente grigio e per il pigmento giallo diffuso nel substrato.Habitat: isole inglesi.

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Zusammenfassung Die Verfasser studieren einen Stamm von Actinomyceten, den sie alsActinomyces Scabies identifizieren. In diesem Zusammenhang stellen sie die diagnostischen Merkmale dieser Art fest, bei kritischer Berücksichtigung der Literatur. Sie stellen fest, dass die unter diesem Namen gesammelten Stämme in den verschiedenen Mycoteken oft verschiedene Merkmale besitzen.Der Grund dieses Missverständnisses liegt, nach den Verfassern, in der pathogenischen Beziehung, i.c., dass man die in den vom Schorf befallenen Knollen die Krankheit als von einer einzigen Art verursacht annehmen will.Nachdem sie die Merkmale der Art festgelegt haben, geben sie die folgende Classification: Actinomyces scabies (Thaxter 1890)Gussov (1914), verbessert von Waksman (1919)=Oospora scabies Thaxter (1890).Diagnose: sieh Seite 273.Habitat: in Nord America ziemlich verbreitet und in Europa das erste Mal vonSpalla undComaschi aus schorfigen Knollen aus Deutschland isoliert. Actinomyces scabies Varietätanglica Baldacci eBoncompagni=Actinomyces scabies Thaxter verbessert vonMillard undBurr.Er unterscheidet sich von der Art durch ein graues vegetatives Mycel, durch ein sehr deutliches graues Luftmycel und durch ein gelbes im Substrat verbreitetes Pigment.Habitat: in England.

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I precedenti Contributi sono stati integralmente pubblicati sugli Atti dell'Istituto Botanico di Pavia e, in riassunto, su questa Rivista.     

Taxonomy and phylogeny of the tribePlucheeae (Asteraceae)

The tribePlucheeae (Benth.)A. Anderb., has been analysed cladistically by means of a computerized parsimony program (Hennig 86), using theArctotideae as outgroup. The results of the analysis are presented in a consensus tree and one cladogram. Four major monophyletic subgroups can be recognized: TheColeocoma group (3 genera), thePterocaulon group (3 genera), theLaggera group (6 genera), and thePluchea group (12 genera). All recognized genera are described and most genera are supplied with taxonomical notes including comments on their taxonomic status. Genera such asBlumea, Pluchea, andEpaltes are demonstrated to be unnatural assemblages.Monarrhenus andTessaria are both closely related to thePluchea complex. The old generic nameLitogyne Harv. has been taken up for one species ofEpaltes, the genusRhodogeron is reduced to a synonym ofSachsia, and the following new combinations are made;Litogyne gariepina (DC.)A. Anderb., andSachsia coronopifolia (Griseb.)A. Anderb.     

Zwei Ausgangskaryotypen bei der Gräser-GattungAira L.

Zusammenfassung Für die Arten der GattungAira L. werden die folgenden Chromosomenzahlen mitgeteilt:A. capillaris Host, 2n=14;A. caryophyllea L., 2n=14 und 28;A. multiculmis Dum., 2n=28;A. praecox L., 2n=14;A. tenorii Guss., 2n=14;A. provincialis Jord., 2n=28 undA. cupaniana Guss., 2n=14. Die Zahlen der letzten drei Arten werden zum erstenmal veröffentlicht.Bei den diploiden Arten kommen zwei unterschiedliche Karyotypen vor: A ist verbreitet, B aufA. praecox beschränkt. Der Karyotyp der tetraploidenA. caryophyllea setzt sich aus A+B zusammen, während es sich beiA. multiculmis um eine Verdoppelung von A handelt. — Über die Verbreitung der Arten und Cytotypen werden einige Hinweise gegeben.

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Two basic karyotypes in the grass genusAira L.

Summary Chromosome numbers of all known species of the genusAira L. are counted:A. capillaris Host, 2n=14;A. caryophyllea L., 2n=14 and 28;A. multiculmis Dum., 2n=28;A. praecox L., 2n=14;A. tenorii Guss., 2n=14;A. provincialis Jord., 2n=28 andA. cupaniana Guss., 2n=14. The numbers of the last three species are published for the first time. Two different karyotypes are found amongst the diploids: A is widespread, B limited toA. praecox. The karyotype of the tetraploidA. caryophyllea can be described as AB, that ofA. multiculmis as AA. — Remarks on the distribution of the species and cytotypes are added.

     

Stimulatory effect of glycine betaine on l-lysine fermentation

Summary The growth rate, sugar consumption rate, and production rate of an l-lysine producing Brevibacterium lactofermentum mutant were stimulated by addition of exogenous glycine betaine. Glycine betaine stimulated the growth rate especially in media of inhibitory osmotic stress, and the stimulation was independent of any specific solute. Therefore growth stimulation by glycine betaine was considered to be an osmoprotective effect. A strong enhancement of the sugar consumption rate and the l-lysine production rate was observed even with resting cells under osmotic stress as well as in a fermentation with growing cells. These data indicated that the osmoprotective effects of glycine betaine on l-lysine production can be independent of protein synthesis. Offprint requests to: Yoshio Kawahara     

Enzymatic preparation of d-p -trimethylsilylphenylalanine

In this paper we report on the enzymatic preparation of d-p-trimethylsilylphenylalanine (d-TMS-Phe). First, dl-5-(p-trimethylsilylphenylmethyl)hydantoin␣(dl-TMS-Phe-Hyd) was synthesized chemically and subjected to bacterial hydrolysis to obtain N-carbamoyl-d-p-trimethylsilylphenylalanine (C-d-TMS-Phe), but no strains examined showed sufficient hydantoinase activity on this compound. However, Blastobacter sp. A17p-4, which is known to produce N-carbamoyl-d-amino acid amidohydrolase (DCase), was found to be able to hydrolyze C-dl-TMS-Phe prepared chemically from the hydantoin. When C-dl-TMS-Phe was hydrolyzed with cells of Blastobacter sp. A17p-4, its optical purity was low because N-carbamoyl-l-amino acid amidohydrolase (LCase) coexisted in the cells. DCase and LCase in the cell-free extract of Blastobacter sp. A17p-4 could be separated by DEAE-Sephacel column chromatography. The optimum pH for the hydrolysis of C-dl-TMS-Phe by the partially purified DCase was 8.0 and addition of 2.5 % N,N-dimethylformamide was effective in raising the substrate concentration without inactivation of DCase. Under the optimized conditions, highly optically pure (98 % enantiomeric excess) d-TMS-Phe could be obtained from C-dl-TMS-Phe with partially purified DCase. Received: 12 July 1996 / Received revision: 11 September 1996 / Accepted: 2 November 1996     

Novos Deuteromycetes da micogeografia intercontinental

Summary This paper deals with some interesting new fungi which are described as a contribution for the development of the Mycogeography.Arbuscula Batista &Peres n. gen. is studied as a member of the family Stilbaceae havingA. eugeniae Batista &Peres n. sp. on leaves ofEugenia jambolana Lam. from Lucknow, India, as the type species. Another new genus isEllisia Batista &Peres, typified byE. ingae Batista &Peres n. sp. on leaves ofInga fagifolia (L.)Willd. from Manaus. Amazonas, Brazil. Two new binomia are:Eriomycopsis paraensis Batista &Peres n. sp. on leaves ofBertholletia excelsa H.B.K. from Belem, State of Pará, Brazil,Sporidesmium americanense Batista &Peres n. sp. on leaves ofQuercus wislizenii A.D.C., from U.S.A. andSporidesmium cookei (Hughes)W. B. Ellis, on leaves of Lauraceae, from Manaus.

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Zusammenfassung Diese Arbeit bringt einige interessante neue Pilze, welche als Beitrag zur Entwicklung der Mycogeographie beschrieben werden.Arbuscula Batista &Peres n. gen. wurde studiert als ein Mitglied der Familie Stilbaceae, dessen TypusA. eugeniae Batista &Peres n. sp. sich auf Blättern vonEugenia jambolana Lam. aus Lucknow, Indien, befand. Eine andere neue Gattung istEllisia Batista &Peres, typiziert durchE. ingae Batista &Peres n. sp. auf Blättern vonInga fagifolia (L.)Willd. aus Manaus, Amazonas, Brasilien. Zwei neue Binome sind:Eriomycopsis paraensis Batista &Peres n. sp. auf Blättern vonBertholletia excelsa H.B.K. aus Belém, Staat Pará, Brasilien,Sporidesmium americanense Batista &Peres n. sp. auf Blättern vonQuercus wislizenii A.D.C. aus U.S.A. undSporidesmium cookei (Hughes)W. B. Ellis, auf Blättern von Lauraceae, aus Manaus, Amazonas.

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Dedicado à Mrmória do Prof.Raffaele Ciferri     

Ein Endemit Zyperns:Asperula cypria (Rubiaceae) und seine Verwandtschaft

Galium suberosum Sibth. etSm. belongs toAsperula sect.Thliphthisa; the taxonomic transfer and a new name are therefore necessary:Asperula cypria Ehrend. is endemic on the isle of Cyprus. Its closest affinities are withA. antalyensis Ehrend. in S.W. Anatolia.