Beta-glucan synthesis in Bradyrhizobium japonicum: characterization of a new locus (ndvC) influencing beta-(1-->6) linkages.

k9 killer toxin from Hansenula mrakii was used to select a number of resistant mutants from Saccharomyces cerevisiae. Preliminary biochemical and genetic studies showed that some of them acquired structural defects in the cell wall. One of these mutants, the knr4-1 mutant, displays a number of cell wall defects, including osmotic sensitivity; sensitivity to cercosporamide, a known antifungal agent; and resistance to Zymolyase, a (1,3)-beta-glucanase. We report here the isolation and analysis of the KNR4 gene. DNA sequence analysis revealed an uninterrupted open reading frame which contains five potential start codons. The longest coding template encodes a protein of 505 amino acids with a calculated molecular mass of 57,044 Da. A data base search revealed 100% identity with a nuclear protein, SMI1p. Disruption of the KNR4 locus does not result in cell death; however, it leads to reduced levels of both (1,3)-beta-glucan synthase activity and (1,3)-beta-glucan content in the cell wall. The gene was mapped to the right arm of chromosome VII.

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Characterization of the yeast (1-->6)-beta-glucan biosynthetic components, Kre6p and Skn1p, and genetic interactions between the PKC1 pathway and extracellular matrix assembly.

The Journal of Cell Biology ◽

10.1083/jcb.127.2.567 ◽

1994 ◽

Vol 127 (2) ◽

pp. 567-579 ◽

Cited By ~ 88

Author(s):

T Roemer ◽

G Paravicini ◽

M A Payton ◽

H Bussey

Keyword(s):

Cell Wall ◽

Beta Glucan ◽

Homologous Proteins ◽

Synthetic Lethal ◽

Cell Wall Assembly ◽

Cell Biol ◽

Synthetic Lethal Interactions ◽

Glucan Synthesis ◽

Wall Assembly

A characterization of the S. cerevisiae KRE6 and SKN1 gene products extends previous genetic studies on their role in (1-->6)-beta-glucan biosynthesis (Roemer, T., and H. Bussey. 1991. Yeast beta-glucan synthesis: KRE6 encodes a predicted type II membrane protein required for glucan synthesis in vivo and for glucan synthase activity in vitro. Proc. Natl. Acad. Sci. USA. 88:11295-11299; Roemer, T., S. Delaney, and H. Bussey. 1993. SKN1 and KRE6 define a pair of functional homologs encoding putative membrane proteins involved in beta-glucan synthesis. Mol. Cell. Biol. 13:4039-4048). KRE6 and SKN1 are predicted to encode homologous proteins that participate in assembly of the cell wall polymer (1-->6)-beta-glucan. KRE6 and SKN1 encode phosphorylated integral-membrane glycoproteins, with Kre6p likely localized within a Golgi subcompartment. Deletion of both these genes is shown to result in a dramatic disorganization of cell wall ultrastructure. Consistent with their direct role in the assembly of this polymer, both Kre6p and Skn1p possess COOH-terminal domains with significant sequence similarity to two recently identified glucan-binding proteins. Deletion of the yeast protein kinase C homolog, PKC1, leads to a lysis defect (Levin, D. E., and E. Bartlett-Heubusch. 1992. Mutants in the S. cerevisiae PKC1 gene display a cell cycle-specific osmotic stability defect. J. Cell Biol. 116:1221-1229). Kre6p when even mildly overproduced, can suppress this pkc1 lysis defect. When mutated, several KRE pathway genes and members of the PKC1-mediated MAP kinase pathway have synthetic lethal interactions as double mutants. These suppression and synthetic lethal interactions, as well as reduced beta-glucan and mannan levels in the pkc1 null wall, support a role for the PKC1 pathway functioning in cell wall assembly. PKC1 potentially participates in cell wall assembly by regulating the synthesis of cell wall components, including (1-->6)-beta-glucan.

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Cloning and characterization of KNR4, a yeast gene involved in (1,3)-beta-glucan synthesis.

Molecular and Cellular Biology ◽

10.1128/mcb.14.2.1017 ◽

1994 ◽

Vol 14 (2) ◽

pp. 1017-1025 ◽

Cited By ~ 61

Author(s):

Z Hong ◽

P Mann ◽

N H Brown ◽

L E Tran ◽

K J Shaw ◽

...

Keyword(s):

Cell Wall ◽

Killer Toxin ◽

Structural Defects ◽

Beta Glucan ◽

Calculated Molecular Mass ◽

Beta Glucanase ◽

Reading Frame ◽

Glucan Synthesis ◽

The Right

k9 killer toxin from Hansenula mrakii was used to select a number of resistant mutants from Saccharomyces cerevisiae. Preliminary biochemical and genetic studies showed that some of them acquired structural defects in the cell wall. One of these mutants, the knr4-1 mutant, displays a number of cell wall defects, including osmotic sensitivity; sensitivity to cercosporamide, a known antifungal agent; and resistance to Zymolyase, a (1,3)-beta-glucanase. We report here the isolation and analysis of the KNR4 gene. DNA sequence analysis revealed an uninterrupted open reading frame which contains five potential start codons. The longest coding template encodes a protein of 505 amino acids with a calculated molecular mass of 57,044 Da. A data base search revealed 100% identity with a nuclear protein, SMI1p. Disruption of the KNR4 locus does not result in cell death; however, it leads to reduced levels of both (1,3)-beta-glucan synthase activity and (1,3)-beta-glucan content in the cell wall. The gene was mapped to the right arm of chromosome VII.

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Characterization of Bradyrhizobium japonicum serocluster 123 member isolates native to Iowa soils

10.31274/rtd-180813-13115 ◽

1992 ◽

Author(s):

Yousef Aharchi

Keyword(s):

Bradyrhizobium Japonicum

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Carbohydrate binding activities of Bradyrhizobium japonicum. II. Isolation and characterization of a galactose-specific lectin.

The Journal of Cell Biology ◽

10.1083/jcb.111.4.1639 ◽

1990 ◽

Vol 111 (4) ◽

pp. 1639-1643 ◽

Cited By ~ 25

Author(s):

S C Ho ◽

M Schindler ◽

J L Wang

Keyword(s):

Isoelectric Focusing ◽

Bradyrhizobium Japonicum ◽

Affinity Column ◽

Carbohydrate Binding ◽

Binding Affinities ◽

Isolation And Characterization ◽

Relative Binding ◽

Mannose Derivatives ◽

Derivatives Of

Extracts of Bradyrhizobium japonicum were fractionated on Sepharose columns covalently derivatized with lactose. Elution of the material that was specifically bound to the affinity column with lactose yielded a protein of Mr approximately 38,000. Isoelectric focusing of this sample yielded two spots with pI values of 6.4 and 6.8. This protein specifically bound to galactose-containing glycoconjugates, but did not bind either to glucose or mannose. Derivatives of galactose at the C-2 position showed much weaker binding; there was an 18-fold difference in the relative binding affinities of galactose versus N-acetyl-D-galactosamine. These results indicate that we have purified a newly identified carbohydrate-binding protein from Bradyrhizobium japonicum, that can exquisitely distinguish galactose from its derivatives at the C-2 position.

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