Molecular cloning of a phospholipid-cholesterol acyltransferase from Aeromonas hydrophila. Sequence homologies with lecithin-cholesterol acyltransferase and other lipases

Thornton, Julian C.; Sp, Howard; Jt, Buckley

doi:10.1016/0005-2760(88)90026-4

Cited by 48 publications

(44 citation statements)

References 20 publications

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“…Budding yeast, Saccharomyces cerevisiae, has only the ACAT type of enzyme for the synthesis of SE (11). In the bacteria, Aeromonas hydrophila, a secreted phospholipase that is structurally unrelated to animal LCAT, has also been shown to catalyze an LCAT reaction (12). The enzyme(s) responsible for SE formation in plants are unknown so far.…”

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confidence: 99%

Cellular Sterol Ester Synthesis in Plants Is Performed by an Enzyme (Phospholipid:Sterol Acyltransferase) Different from the Yeast and Mammalian Acyl-CoA:Sterol Acyltransferases

Banaś

Carlsson

Huang

et al. 2005

Journal of Biological Chemistry

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A gene encoding a sterol ester-synthesizing enzyme was identified in Arabidopsis. The cDNA of the Arabidopsis gene At1g04010 (AtPSAT) was overexpressed in Arabidopsis behind the cauliflower mosaic virus 35S promoter. Microsomal membranes from the leaves of overexpresser lines catalyzed the transacylation of acyl groups from phosphatidylethanolamine to sterols. This activity correlated with the expression level of the AtPSAT gene, thus demonstrating that this gene encodes a phospholipid:sterol acyltransferase (PSAT). Properties of the AtPSAT were examined in microsomal fractions from the tissues of an overexpresser. The enzyme did not utilize neutral lipids, had the highest activity with phosphatidylethanolamine, had a 5-fold preference for the sn-2 position, and utilized both saturated and unsaturated fatty acids. Various sterols and sterol intermediates, including triterpenic precursors, were acylated by the PSAT, whereas other triterpenes were not. Sterol selectivity studies showed that the enzyme is activated by end product sterols and that sterol intermediates are preferentially acylated by the activated enzyme. This indicates that PSAT both regulates the pool of free sterols as well as limits the amount of free sterol intermediates in the membranes. Two T-DNA insertion mutants in the AtPSAT gene, with strongly reduced (but still measurable) levels of sterol esters in their tissues, had no detectable PSAT activity in the microsomal fractions, suggesting that Arabidopsis possess other enzyme(s) capable of acylating sterols. The AtPSAT is the only intracellular enzyme found so far that catalyzes an acyl-CoA-independent sterol ester formation. Thus, PSAT has a similar physiological function in plant cells as the unrelated acyl-CoA:sterol acyltransferase has in animal cells.Sterols are components of all eukaryotic membranes and are important regulators of membrane fluidity and thus membrane properties and functions (1, 2). Because acylated sterols or sterol esters (SE) 2 cannot participate in the bilayer of the membranes, the acylation of sterols is believed to play a crucial role in maintaining free sterol homeostasis in the cell membranes (3-7). In other words, sterol esters are generally thought to constitute a storage pool of sterols when those are present in amounts greater than immediately required for the cells. For instance, accumulation of sterol esters has been described during seed maturation, senescence, or when plant cell cultures reach stationary phase, as well as in a mutant line overproducing sterols (4).Although no plant gene involved in sterol esterification had been isolated yet, the animal and yeast counterparts have been well characterized. Two types of enzymes are responsible for the formation of SE in animals as follows: acyl-CoA:cholesterol acyltransferase (ACAT), which catalyzes an acyl-CoA-dependent acylation (7), and lecithin:cholesterol acyltransferase (LCAT), which catalyzes transacylation of acyl groups from phospholipids to sterols (8).ACAT is membrane-bound and has sequence homolo...

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confidence: 99%

Cellular Sterol Ester Synthesis in Plants Is Performed by an Enzyme (Phospholipid:Sterol Acyltransferase) Different from the Yeast and Mammalian Acyl-CoA:Sterol Acyltransferases

Banaś

Carlsson

Huang

et al. 2005

Journal of Biological Chemistry

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“…hydrophila MCC-1 a-amylase gene was 63 mol YO, whilst that of the third position of the codons was higher (83 mol%). These values are nearly identical to those for glycerophospholipid<holesterol acyltransferase (GCAT) [62 and 82 mol%, respectively (Thornton et al, 1988)], amylase [62 and 82 mol YO, respectively (Gobius & Pemberton, 1988)l and deoxyribonuclease (DNAase) [63 and 87 mol%, respectively (Chang et al, 1992)], and are a little higher than that for aerolysin [58 and 72 mol%, respectively (Howard et al, 1987)] from this species. The codon usage frequence in the a-amylase gene is similar to the pattern for the aforementioned genes of this species (data not shown).…”

Section: Codon Usage Patterns Of the A-amylasementioning

confidence: 55%

“…hydrophila, this organism has been chosen for the study of secretion mechanisms. Previous studies have reported that when aerolysin, acyltransferase, protease and amylase structural genes are expressed in Escherichia coli, these exoproteins are secreted only to the periplasm, indicating that specific functions, absent in E. coli, were necessary for the second step of transfer through the outer membrane and release into the medium (Chakraborty et al, 1986;Gobius & Pemberton, 1988;Howard & Buckley, 1986;Rivero et al, 1990;Thornton et al, 1988). The need for other gene products is supported by the observation that pleiotropic export mutants of A .…”

Section: Introductionmentioning

confidence: 95%

Cloning and nucleotide sequence of an extracellular -amylase gene from Aeromonas hydrophila MCC-1

Chang

Chang²,

Chen³

1993

Journal of General Microbiology

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A gene encoding the extracellular a-amylase of Aevomonas hydrophila MCC-1 was cloned and expressed using its own promoter on the recombinant plasmid pCA101. Subcellular fractionation of Eschevichia coli JA221 carrying pCAlOl revealed that approximately 60 YO of the amylase activity was localized in the periplasmic space. The extracellular amylase was purified to homogeneity, identified as an a-type and its amino-terminal sequence was determined. Nucleotide sequence analysis predicted a 443 amino acid ORF and 24 amino acids at the amino terminus of the sequence that are not found in the secreted protein. This 24 amino acid sequence has many of the characteristics common to known signal peptides. The predicted amino acid sequence has considerable similarity with mammalian, invertebrate and Streptomycete a-amylases. Most of the amino acid residues that are involved in catalytic activity, substrate binding and calcium binding in several a-amylases were also present in A . hydrophila a-amylase at the corresponding positions.

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“…Although the limited size of the dbEST sequences prohibits extensive sequence alignment, it was apparent that the great majority of these translated dbEST sequences also contained a PYG motif shortly after Block I. In subsequent database searches we discovered that the simple, single motif (10,20)PY, a modified Block I motif, is absolutely specific for the translated cDNAs from the dbEST and the two plant proteins previously identified. This indicates that the database search results are significant and that these sequences are indeed related.…”

Section: Resultsmentioning

confidence: 95%

“…proteins in this group are known to possess lipolytic activity. These include a lipase/acyltransferase from Aeromonas hydrophila [10], a hemolysin from Vibrio parahaemolyticus [11] and a lipase from Xenorhabdus luminescens [12]. In addition, we have already used our motif scheme to identify the catalytic triad of the Aeromonas hydrophila enzyme [13].…”

Section: Introductionmentioning

confidence: 99%

A new family of lipolytic plant enzymes with members in rice, arabidopsis and maize

et al. 1995

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We have noted a striking similarity between the sequences of proteins in a novel family of lipases we recently reported [Upton, C. and Buckley, J.T. (1995) Trends Biol. Sci. 20, 178-9] and more than 120 sequences from the database of Expressed Sequence Tags (dbEST) which correspond to at least 30 unique genes from arabidopsis, rice and maize. A cDNA (Arab-l) corresponding to one of these sequences was isolated, sequenced and translated. There was significant similarity to sequences in the new 5pase family over the entire open reading frame of Arab-1 and when expressed in E. coil, the gene product was lipolytic. Arab-1 and genes for some of the other plant proteins appear to be differentially expressed. They may play a role in the regulation of lipid metabolism during plant development.

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Molecular cloning of a phospholipid-cholesterol acyltransferase from Aeromonas hydrophila. Sequence homologies with lecithin-cholesterol acyltransferase and other lipases

Cited by 48 publications

References 20 publications

Cellular Sterol Ester Synthesis in Plants Is Performed by an Enzyme (Phospholipid:Sterol Acyltransferase) Different from the Yeast and Mammalian Acyl-CoA:Sterol Acyltransferases

Cellular Sterol Ester Synthesis in Plants Is Performed by an Enzyme (Phospholipid:Sterol Acyltransferase) Different from the Yeast and Mammalian Acyl-CoA:Sterol Acyltransferases

Cloning and nucleotide sequence of an extracellular -amylase gene from Aeromonas hydrophila MCC-1

A new family of lipolytic plant enzymes with members in rice, arabidopsis and maize

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