Análisis del genoma de Pycnoporus cinnabarinus mediante electroforesis de campo pulsado

Boletín Micológico ◽

10.22370/bolmicol.1992.7.0.1192 ◽

2018 ◽

Vol 7 ◽

pp. 31

Author(s):

Víctor Cifuentes ◽

Claudio Martínez

Keyword(s):

Saccharomyces Cerevisiae ◽

Pycnoporus Cinnabarinus ◽

Schwanniomyces Occidentalis

En el presente trabajo se describe la separación óptima de moléculas de DNA cromosómico intacto del hongo filamentoso Pycnoporus cinnabarinus, mediante eléctroforesis de campo pulsado usando un sistema de contorno de campo eléctrico homogéneo (CHEF), con un electrodo hexagonal.Se estudió una serie de condiciones experimentales en las cuales se hizo variar la duración del intervalo de tiempo en cada dirección del campo y el tiempo total de la corrida.De esta manera fue posible separar el DNA cromosómico de P. cinnabarinus en cuatro bandas que, dependiendo e las condiciones de electroforesis,pueden ser resueltas por lo menos en siete bandas. Utilizando los patrones de migración del DNA cromosómico de Saccharomyces cerevisiae y Schwanniomyces occidentalis comoestándar, estimamos que los tamaños moleculares del DNA cromosómico de P. cinnabarinus oscilan entre 1.9 y 2.5 megabases (Mb).

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Expression of the Schwanniomyces occidentalis SWA2 amylase in Saccharomyces cerevisiae: role of N-glycosylation on activity, stability and secretion

Biochemical Journal ◽

10.1042/bj3290065 ◽

1998 ◽

Vol 329 (1) ◽

pp. 65-71 ◽

Cited By ~ 36

Author(s):

Esther YÁÑEZ ◽

A. Teresa CARMONA ◽

Mercedes TIEMBLO ◽

Antonio JIMÉNEZ ◽

María FERNÁNDEZ-LOBATO

Keyword(s):

Saccharomyces Cerevisiae ◽

Catalytic Efficiency ◽

Extracellular Enzyme ◽

Site Directed Mutagenesis ◽

Activity Levels ◽

Wild Type ◽

Schwanniomyces Occidentalis ◽

Type Protein ◽

Wild Type Protein

The role of N-linked glycosylation on the biological activity of Schwanniomyces occidentalis SWA2 α-amylase, as expressed in Saccharomyces cerevisiae, was analysed by site-directed mutagenesis of the two potential N-glycosylation sites, Asn-134 and Asn-229. These residues were replaced by Ala or Gly individually or in various combinations and the effects on the activity, secretion and thermal stability of the enzyme were studied. Any Asn-229 substitution caused a drastic decrease in activity levels of the extracellular enzyme. In contrast, substitutions of Asn-134 had little or no effect. The use of antibodies showed that α-amylase was secreted in all the mutants tested, although those containing substitutions at Asn-229 seemed to have a lower rate of synthesis and/or higher degradation than the wild-type strain. α-Amylases with substitution at Asn-229 had a 2 kDa lower molecular mass than the wild-type protein, as did the wild-type protein itself after treatment with endoglycosidase F. These findings indicate that Asn-229 is the single glycosylated residue in SWA2. Thermostability analysis of both purified wild-type (T50 = 50 °C, where T50 is the temperature resulting in 50% loss of activity) and mutant enzymes indicated that removal of carbohydrate from the 229 position results in a decrease of approx. 3 °C in the T50 of the enzyme. The Gly-229 mutation does not change the apparent affinity of the enzyme for starch (Km) but decreases to 1/22 its apparent catalytic efficiency (kcat/Km). These results therefore indicate that glycosylation at the 229 position has an important role in the extracellular activity levels, kinetics and stability of the Sw. occidentalis SWA2 α-amylase in both its wild-type and mutant forms.

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Fermentation of grain sorghum starch by co-cultivation of schwanniomyces occidentalis and saccharomyces cerevisiae

Bioresource Technology ◽

10.1016/0960-8524(92)90084-b ◽

1992 ◽

Vol 42 (1) ◽

pp. 27-31 ◽

Cited By ~ 13

Author(s):

C.H. Horn ◽

J.C. du Preez ◽

S.G. Kilian

Keyword(s):

Saccharomyces Cerevisiae ◽

Grain Sorghum ◽

Schwanniomyces Occidentalis

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Cloning of the Schwanniomyces occidentalis glucoamylase gene (GAM1) and its expression in Saccharomyces cerevisiae

Gene ◽

10.1016/0378-1119(90)90421-m ◽

1990 ◽

Vol 95 (1) ◽

pp. 111-121 ◽

Cited By ~ 50

Author(s):

R.Jürgen Dohmen ◽

Alexander W.M. Strasser ◽

Ulrike M. Dahlems ◽

Cornelis P. Hollenberg

Keyword(s):

Saccharomyces Cerevisiae ◽

Schwanniomyces Occidentalis ◽

Glucoamylase Gene

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Cloning and Expression of Two Genes Coding for Sodium Pumps in the Salt-Tolerant Yeast Debaryomyces hansenii

Journal of Bacteriology ◽

10.1128/jb.183.10.3251-3255.2001 ◽

2001 ◽

Vol 183 (10) ◽

pp. 3251-3255 ◽

Cited By ~ 65

Author(s):

Anabel Almagro ◽

Catarina Prista ◽

Begoña Benito ◽

Maria C. Loureiro-Dias ◽

José Ramos

Keyword(s):

Saccharomyces Cerevisiae ◽

Debaryomyces Hansenii ◽

Cloning And Expression ◽

High Homology ◽

Salt Tolerant ◽

Schwanniomyces Occidentalis ◽

High Ph ◽

Sodium Pumps ◽

Genes Encoding

ABSTRACT Two genes encoding Na+-ATPases from Debaryomyces hansenii were cloned and sequenced. The genes, designatedENA1 from D. hansenii (DhENA1) andDhENA2, exhibited high homology with the corresponding genes from Schwanniomyces occidentalis. DhENA1 was expressed in the presence of high Na+ concentrations, while the expression of DhENA2 also required high pH. A mutant ofSaccharomyces cerevisiae lacking the Na+ efflux systems and sensitive to Na+, when transformed withDhENA1 or DhENA2, recovered Na+tolerance and also the ability to extrude Na+.

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