scholarly journals Structural and functional conservation between yeast and human 3-hydroxy-3-methylglutaryl coenzyme A reductases, the rate-limiting enzyme of sterol biosynthesis.

1988 ◽  
Vol 8 (9) ◽  
pp. 3797-3808 ◽  
Author(s):  
M E Basson ◽  
M Thorsness ◽  
J Finer-Moore ◽  
R M Stroud ◽  
J Rine
Keyword(s):  
Coenzyme A ◽  
Sterol Biosynthesis ◽  
New Drugs ◽  
Yeast Cells ◽  
Cdna Clones ◽  
Hmg Coa Reductase ◽  
Functional Conservation ◽  
Mammalian Enzyme ◽  
Coa Reductase ◽  
Rate Limiting

The pathway of sterol biosynthesis is highly conserved in all eucaryotic cells. We demonstrated structural and functional conservation of the rate-limiting enzyme of the mammalian pathway, 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMG-CoA reductase), between the yeast Saccharomyces cerevisiae and humans. The amino acid sequence of the two yeast HMG-CoA reductase isozymes was deduced from DNA sequence analysis of the HMG1 and HMG2 genes. Extensive sequence similarity existed between the region of the mammalian enzyme encoding the active site and the corresponding region of the two yeast isozymes. Moreover, each of the yeast isozymes, like the mammalian enzyme, contained seven potential membrane-spanning domains in the NH2-terminal region of the protein. Expression of cDNA clones encoding either hamster or human HMG-CoA reductase rescued the viability of hmg1 hmg2 yeast cells lacking this enzyme. Thus, mammalian HMG-CoA reductase can provide sufficient catalytic function to replace both yeast isozymes in vivo. The availability of yeast cells whose growth depends on human HMG-CoA reductase may provide a microbial screen to identify new drugs that can modulate cholesterol biosynthesis.

Structural and functional conservation between yeast and human 3-hydroxy-3-methylglutaryl coenzyme A reductases, the rate-limiting enzyme of sterol biosynthesis

1988 ◽  
Vol 8 (9) ◽  
pp. 3797-3808
Author(s):  
M E Basson ◽  
M Thorsness ◽  
J Finer-Moore ◽  
R M Stroud ◽  
J Rine
Keyword(s):  
Coenzyme A ◽  
Sterol Biosynthesis ◽  
New Drugs ◽  
Yeast Cells ◽  
Cdna Clones ◽  
Hmg Coa Reductase ◽  
Functional Conservation ◽  
Mammalian Enzyme ◽  
Coa Reductase ◽  
Rate Limiting

The pathway of sterol biosynthesis is highly conserved in all eucaryotic cells. We demonstrated structural and functional conservation of the rate-limiting enzyme of the mammalian pathway, 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMG-CoA reductase), between the yeast Saccharomyces cerevisiae and humans. The amino acid sequence of the two yeast HMG-CoA reductase isozymes was deduced from DNA sequence analysis of the HMG1 and HMG2 genes. Extensive sequence similarity existed between the region of the mammalian enzyme encoding the active site and the corresponding region of the two yeast isozymes. Moreover, each of the yeast isozymes, like the mammalian enzyme, contained seven potential membrane-spanning domains in the NH2-terminal region of the protein. Expression of cDNA clones encoding either hamster or human HMG-CoA reductase rescued the viability of hmg1 hmg2 yeast cells lacking this enzyme. Thus, mammalian HMG-CoA reductase can provide sufficient catalytic function to replace both yeast isozymes in vivo. The availability of yeast cells whose growth depends on human HMG-CoA reductase may provide a microbial screen to identify new drugs that can modulate cholesterol biosynthesis.

Genetic and biochemical evaluation of eucaryotic membrane protein topology: multiple transmembrane domains of Saccharomyces cerevisiae 3-hydroxy-3-methylglutaryl coenzyme A reductase

1990 ◽  
Vol 10 (2) ◽  
pp. 672-680
Author(s):  
C Sengstag ◽  
C Stirling ◽  
R Schekman ◽  
J Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Coenzyme A ◽  
Transmembrane Domains ◽  
Protein Domain ◽  
Yeast Cells ◽  
Endoplasmic Reticulum Membrane ◽  
Hmg Coa Reductase ◽  
Coa Reductase ◽  
Histidinol Dehydrogenase

Both 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase isozymes of the yeast Saccharomyces cerevisiae are predicted to contain seven membrane-spanning domains. Previous work had established the utility of the histidinol dehydrogenase protein domain, encoded by HIS4C, as a topologically sensitive monitor that can be used to distinguish between the lumen of the endoplasmic reticulum and the cytoplasm. This study directly tested the structural predictions for HMG-CoA reductase by fusing the HIS4C domain to specific sites in the HMG-CoA reductase isozymes. Yeast cells containing the HMG-CoA reductase-histidinol dehydrogenase fusion proteins grew on histidinol-containing medium if the HIS4C domain was present on the cytoplasmic side of the endoplasmic reticulum membrane but not if the HIS4C domain was targeted to the endoplasmic reticulum lumen. Systematic exchanges of transmembrane domains between the isozymes confirmed that both isozymes had equivalent membrane topologies. In general, deletion of an even number of putative transmembrane domains did not interfere with the topology of the protein, but deletion or duplication of an odd number of transmembrane domains inverted the orientation of the protein. The data confirmed the earlier proposed topology for yeast HMG-CoA reductase, demonstrated that the yeast enzymes are core glycosylated, and provided in vivo evidence that the properties of transmembrane domains were, in part, dependent upon their context within the protein.

scholarly journals Developmental and metabolic regulation of the Drosophila melanogaster 3-hydroxy-3-methylglutaryl coenzyme A reductase.

1988 ◽  
Vol 8 (7) ◽  
pp. 2713-2721 ◽  
Author(s):  
F B Gertler ◽  
C Y Chiu ◽  
L Richter-Mann ◽  
D J Chin
Keyword(s):  
Drosophila Melanogaster ◽  
Metabolic Regulation ◽  
Coenzyme A ◽  
Terminal Region ◽  
Cdna Clones ◽  
Posttranslational Regulation ◽  
Hmg Coa Reductase ◽  
Reading Frame ◽  
Reductase Gene ◽  
Coa Reductase

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in Drosophila melanogaster synthesizes mevalonate for the production of nonsterol isoprenoids, which are essential for growth and differentiation. To understand the regulation and developmental role of HMG CoA reductase, we cloned the D. melanogaster HMG CoA reductase gene. The nucleotide sequence of the Drosophila HMG CoA reductase was determined from genomic and cDNA clones. A 2,748-base-pair open reading frame encoded a polypeptide of 916 amino acids (Mr, 98,165) that was similar to the hamster HMG CoA reductase. The C-terminal region had 56% identical residues and the N-terminal region had 7 potential transmembrane domains with 32 to 60% identical residues. In hamster HMG CoA reductase, the membrane regions were essential for posttranslational regulation. Since the Drosophila enzyme is not regulated by sterols, the strong N-terminal similarity was surprising. Two HMG CoA reductase mRNA transcripts, approximately 3.2 and 4 kilobases, were differentially expressed throughout Drosophila development. Mevalonate-fed Schneider cells showed a parallel reduction of both enzyme activity and abundance of the 4-kilobase mRNA transcript.

scholarly journals Genetic and biochemical evaluation of eucaryotic membrane protein topology: multiple transmembrane domains of Saccharomyces cerevisiae 3-hydroxy-3-methylglutaryl coenzyme A reductase.

1990 ◽  
Vol 10 (2) ◽  
pp. 672-680 ◽  
Author(s):  
C Sengstag ◽  
C Stirling ◽  
R Schekman ◽  
J Rine
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Endoplasmic Reticulum ◽  
Coenzyme A ◽  
Transmembrane Domains ◽  
Protein Domain ◽  
Yeast Cells ◽  
Endoplasmic Reticulum Membrane ◽  
Hmg Coa Reductase ◽  
Coa Reductase ◽  
Histidinol Dehydrogenase

Both 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase isozymes of the yeast Saccharomyces cerevisiae are predicted to contain seven membrane-spanning domains. Previous work had established the utility of the histidinol dehydrogenase protein domain, encoded by HIS4C, as a topologically sensitive monitor that can be used to distinguish between the lumen of the endoplasmic reticulum and the cytoplasm. This study directly tested the structural predictions for HMG-CoA reductase by fusing the HIS4C domain to specific sites in the HMG-CoA reductase isozymes. Yeast cells containing the HMG-CoA reductase-histidinol dehydrogenase fusion proteins grew on histidinol-containing medium if the HIS4C domain was present on the cytoplasmic side of the endoplasmic reticulum membrane but not if the HIS4C domain was targeted to the endoplasmic reticulum lumen. Systematic exchanges of transmembrane domains between the isozymes confirmed that both isozymes had equivalent membrane topologies. In general, deletion of an even number of putative transmembrane domains did not interfere with the topology of the protein, but deletion or duplication of an odd number of transmembrane domains inverted the orientation of the protein. The data confirmed the earlier proposed topology for yeast HMG-CoA reductase, demonstrated that the yeast enzymes are core glycosylated, and provided in vivo evidence that the properties of transmembrane domains were, in part, dependent upon their context within the protein.

Developmental and metabolic regulation of the Drosophila melanogaster 3-hydroxy-3-methylglutaryl coenzyme A reductase

1988 ◽  
Vol 8 (7) ◽  
pp. 2713-2721
Author(s):  
F B Gertler ◽  
C Y Chiu ◽  
L Richter-Mann ◽  
D J Chin
Keyword(s):  
Drosophila Melanogaster ◽  
Metabolic Regulation ◽  
Coenzyme A ◽  
Terminal Region ◽  
Cdna Clones ◽  
Posttranslational Regulation ◽  
Hmg Coa Reductase ◽  
Reading Frame ◽  
Reductase Gene ◽  
Coa Reductase

The enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in Drosophila melanogaster synthesizes mevalonate for the production of nonsterol isoprenoids, which are essential for growth and differentiation. To understand the regulation and developmental role of HMG CoA reductase, we cloned the D. melanogaster HMG CoA reductase gene. The nucleotide sequence of the Drosophila HMG CoA reductase was determined from genomic and cDNA clones. A 2,748-base-pair open reading frame encoded a polypeptide of 916 amino acids (Mr, 98,165) that was similar to the hamster HMG CoA reductase. The C-terminal region had 56% identical residues and the N-terminal region had 7 potential transmembrane domains with 32 to 60% identical residues. In hamster HMG CoA reductase, the membrane regions were essential for posttranslational regulation. Since the Drosophila enzyme is not regulated by sterols, the strong N-terminal similarity was surprising. Two HMG CoA reductase mRNA transcripts, approximately 3.2 and 4 kilobases, were differentially expressed throughout Drosophila development. Mevalonate-fed Schneider cells showed a parallel reduction of both enzyme activity and abundance of the 4-kilobase mRNA transcript.

scholarly journals Dietary modification of the activation state of intestinal 3-hydroxy-3-methylglutaryl coenzyme a (HMG-CoA) reductase.

1984 ◽  
Vol 48 (11) ◽  
pp. 2745-2751
Author(s):  
Hirosuke OKU ◽  
Akira MORITA ◽  
Takashi IDE ◽  
Michihiro SUGANO
Keyword(s):  
Coenzyme A ◽  
Dietary Modification ◽  
Hmg Coa Reductase ◽  
Coa Reductase

scholarly journals The sterol-responsive RNF145 E3 ubiquitin ligase mediates the degradation of HMG-CoA reductase together with gp78 and Hrd1

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Sam A Menzies ◽  
Norbert Volkmar ◽  
Dick JH van den Boomen ◽  
Richard T Timms ◽  
Anna S Dickson ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Critical Role ◽  
E3 Ubiquitin Ligase ◽  
E3 Ligase ◽  
Editorial Note ◽  
Hmg Coa Reductase ◽  
Genome Wide ◽  
Cholesterol Biosynthetic Pathway ◽  
Coa Reductase ◽  
Rate Limiting

Mammalian HMG-CoA reductase (HMGCR), the rate-limiting enzyme of the cholesterol biosynthetic pathway and the therapeutic target of statins, is post-transcriptionally regulated by sterol-accelerated degradation. Under cholesterol-replete conditions, HMGCR is ubiquitinated and degraded, but the identity of the E3 ubiquitin ligase(s) responsible for mammalian HMGCR turnover remains controversial. Using systematic, unbiased CRISPR/Cas9 genome-wide screens with a sterol-sensitive endogenous HMGCR reporter, we comprehensively map the E3 ligase landscape required for sterol-accelerated HMGCR degradation. We find that RNF145 and gp78 independently co-ordinate HMGCR ubiquitination and degradation. RNF145, a sterol-responsive ER-resident E3 ligase, is unstable but accumulates following sterol depletion. Sterol addition triggers RNF145 recruitment to HMGCR via Insigs, promoting HMGCR ubiquitination and proteasome-mediated degradation. In the absence of both RNF145 and gp78, Hrd1, a third UBE2G2-dependent E3 ligase, partially regulates HMGCR activity. Our findings reveal a critical role for the sterol-responsive RNF145 in HMGCR regulation and elucidate the complexity of sterol-accelerated HMGCR degradation.Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (<xref ref-type="decision-letter" rid="SA1">see decision letter</xref>).

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