scholarly journals Identification and functional characterization of the HpALG11 and the HpRFT1 genes involved in N-linked glycosylation in the methylotrophic yeast Hansenula polymorpha

Glycobiology ◽  
2010 ◽  
Vol 20 (12) ◽  
pp. 1665-1674 ◽  
Author(s):  
Haolei Song ◽  
Weidong Qian ◽  
Hui Wang ◽  
Bingsheng Qiu
Keyword(s):  
Hansenula Polymorpha ◽  
Functional Characterization ◽  
Methylotrophic Yeast

scholarly journals The Hansenula polymorpha PER8 gene encodes a novel peroxisomal integral membrane protein involved in proliferation.

1995 ◽  
Vol 128 (3) ◽  
pp. 307-319 ◽  
Author(s):  
X Tan ◽  
H R Waterham ◽  
M Veenhuis ◽  
J M Cregg
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Hansenula Polymorpha ◽  
Methylotrophic Yeast ◽  
Integral Membrane Protein ◽  
Peroxisome Biogenesis ◽  
Primary Sequence ◽  
Zinc Finger Motifs ◽  
Molecular Machinery

We previously described the isolation of mutants of the methylotrophic yeast Hansenula polymorpha that are defective in peroxisome biogenesis. Here, we describe the characterization of one of these mutants, per8, and the cloning of the PER8 gene. In either methanol or methylamine medium, conditions that normally induce the organelles, per8 cells contain no peroxisome-like structures and peroxisomal enzymes are located in the cytosol. The sequence of PER8 predicts that its product (Per8p) is a novel polypeptide of 34 kD, and antibodies against Per8p recognize a protein of 31 kD. Analysis of the primary sequence of Per8p revealed a 39-amino-acid cysteine-rich segment with similarity to the C3HC4 family of zinc-finger motifs. Overexpression of PER8 results in a markedly enhanced increase in peroxisome numbers. We show that Per8p is an integral membrane protein of the peroxisome and that it is concentrated in the membranes of newly formed organelles. We propose that Per8p is a component of the molecular machinery that controls the proliferation of this organelle.

Cloning and biochemical characterization of hexokinase from the methylotrophic yeast Hansenula polymorpha

2003 ◽  
Vol 44 (5) ◽  
pp. 268-276 ◽  
Author(s):  
Helen Karp ◽  
Aiki J�rviste ◽  
Thomas M. Kriegel ◽  
Tiina Alam�e
Keyword(s):  
Biochemical Characterization ◽  
Hansenula Polymorpha ◽  
Methylotrophic Yeast

Cloning and characterization of the gene encoding a repressible acid phosphatase ( PHO1 ) from the methylotrophic yeast Hansenula polymorpha

1998 ◽  
Vol 50 (1) ◽  
pp. 77-84 ◽  
Author(s):  
A. Phongdara ◽  
A. Merckelbach ◽  
P. Keup ◽  
G. Gellissen ◽  
C. P. Hollenberg
Keyword(s):  
Acid Phosphatase ◽  
Hansenula Polymorpha ◽  
Methylotrophic Yeast ◽  
Gene Encoding

scholarly journals Yeast Cell-Based Transport Assay for the Functional Characterization of Human 4F2hc-LAT1 and ‐LAT2, and LAT1 and LAT2 Substrates and Inhibitors

2021 ◽  
Vol 8 ◽  
Author(s):  
Satish Kantipudi ◽  
Dimitrios Fotiadis
Keyword(s):  
Amino Acid ◽  
Mammalian Cells ◽  
Functional Characterization ◽  
Methylotrophic Yeast ◽  
Cell Types ◽  
Amino Acid Transporters ◽  
Cell Assays ◽  
Transport Assay ◽  
Compound Screening

In mammalian cells, the L-type amino acid transporters (LATs) LAT1 (SLC7A5) and LAT2 (SLC7A8) form heterodimeric amino acid transporters (HATs) with the ancillary protein 4F2hc and are involved in the cellular uptake of specific amino acids. The HAT 4F2hc-LAT1 is found upregulated in various cancer cell types, while 4F2hc-LAT2 is a transporter for non-cancer cells. Preclinical studies have highlighted that 4F2hc-LAT1 plays an important role in tumor progression representing a valid anticancer target. Consequently, current research is focusing on the development of potent and specific human 4F2hc-LAT1 inhibitors. On the other hand, 4F2hc-LAT2 is emerging as target of other diseases, thus also gaining clinical interest. To determine affinity and specificity of substrates and inhibitors for 4F2hc-LAT1 or 4F2hc-LAT2, robust transport cell assays are indispensable. We have optimized and validated a transport assay using cells of the methylotrophic yeast Pichia pastoris stably overexpressing the human HATs 4F2hc-LAT1 or -LAT2, and the LATs LAT1 or LAT2 alone. The radioligand [3H]L-leucine was used as reporter and the substrates L-leucine, triiodothyronine (T3) and thyroxine (T4) as well as the inhibitors BCH and JPH203 (KYT-0353) for assay validation. Obtained half-maximal inhibitory concentrations also provided new insights, e.g., into the LAT specificity of the potent inhibitor JPH203 and on the potency of the thyroid hormones T3 and T4 to inhibit transport through human 4F2hc-LAT2. The LAT1 and LAT2 assays are of particular interest to determine possible implications and influences of 4F2hc in ligand binding and transport. In summary, the presented assays are valuable for characterization of ligands, e.g., towards 4F2hc-LAT1 specificity, and can also be applied for compound screening. Finally, our established approach and assay would also be applicable to other HATs and LATs of interest.

scholarly journals Isolation and characterization of cold sensitive pex6 mutant of the methylotrophic yeast Hansenula polymorpha

2002 ◽  
Vol 18 (2) ◽  
pp. 131-134 ◽  
Author(s):  
V. Y. Nazarko ◽  
O. D. Pochapinsky ◽  
T. Y. Nazarko ◽  
O. V. Stasyk ◽  
A. A. Sibirny
Keyword(s):  
Hansenula Polymorpha ◽  
Methylotrophic Yeast ◽  
Isolation And Characterization ◽  
Cold Sensitive

scholarly journals A novel Delta12-fatty acid desaturase gene from methylotrophic yeast Pichia pastoris GS115.

2006 ◽  
Vol 53 (4) ◽  
pp. 753-759 ◽  
Author(s):  
D Sh Wei ◽  
M Ch Li ◽  
X X Zhang ◽  
H Zhou ◽  
L J Xing
Keyword(s):  
Fatty Acid ◽  
Functional Characterization ◽  
Fatty Acid Desaturase ◽  
Methylotrophic Yeast ◽  
Pastoris Gs115 ◽  
Desaturase Gene ◽  
Methylotrophic Yeast Pichia Pastoris ◽  
Membrane Bound ◽  
Fatty Acid Desaturase Gene

The methylotrophic yeast Pichia pastoris GS115, a widely used strain in production of various heterologous proteins, especially membrane-bound enzymes, can also produce linoleic and linolenic acids, which indicates the existence of membrane-bound Delta12 and Delta15-fatty acid desaturases. This paper describes the cloning and functional characterization of a novel Delta12-fatty acid desaturase gene from this methylotrophic yeast. The open reading frame of the gene (named Pp-FAD12) is 1263 bp in size and encodes a 420-amino-acid peptide. The deduced Pp-FAD12 protein shows high identity (50-67%) with Delta12-fatty acid desaturases from other fungi. It also shows a high identity (57%) with Delta15-fatty acid desaturase (named Sk-FAD15) from Saccharomyces kluyveri. Expression of Pp-FAD12 in polyunsaturated fatty acids non-producing yeast Saccharomyces cerevisiae demonstrated that its product converted oleic acid (18 : 1) to linoleic acid (18 : 2). This result suggests that Pp-FAD12 encodes a novel Delta12-fatty acid desaturase in P. pastoris GS115. This is the first report about the cloning and functional characterization of Delta12-fatty acid desaturase gene in methylotrophic yeast.

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