東華大學圖書館 |

Language: English

Help

回圖書館首頁

手機版館藏查詢

Back

Switch To: Labeled | MARC Mode | ISBD

Functional characterization of stero...

Guo, Zhongmin.

Linked to FindBook

Google Book

Amazon

博客來

Functional characterization of sterol esterification enzymes in yeast.

Record Type:	Electronic resources : Monograph/item
Title/Author:	Functional characterization of sterol esterification enzymes in yeast./
Author:	Guo, Zhongmin.
Published:	Ann Arbor : ProQuest Dissertations & Theses, : 2000,
Description:	142 p.
Notes:	Source: Dissertations Abstracts International, Volume: 62-03, Section: B.
Contained By:	Dissertations Abstracts International62-03B.
Subject:	Nutrition. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=9970202
ISBN:	9780599751682

Functional characterization of sterol esterification enzymes in yeast.
Guo, Zhongmin.

Functional characterization of sterol esterification enzymes in yeast. - Ann Arbor : ProQuest Dissertations & Theses, 2000 - 142 p.

Source: Dissertations Abstracts International, Volume: 62-03, Section: B.

Thesis (Ph.D.)--Columbia University, 2000.

This item must not be sold to any third party vendors.

In all eukaryotes, free sterol is an essential component of membranes. However, high levels of free sterol can be toxic. Cells maintain sterol homeostasis, in part, by converting excess free sterol to sterol ester. In mammals this reaction is mediated by the enzyme acyl-coenzyme A: cholesterol acyltransferase (ACAT). ACAT activity has been implicated in multiple physiologic processes, and in the pathological development of atherosclerosis. Two yeast ACAT homologs, ARE1 and ARE2 (for ACAT related enzymes 1 and 2, respectively), were identified, disruption of both genes produced a viable yeast cell with no detectable sterol ester. The goal of this study was to investigate the mechanism and function of sterol esterification. enzymes using yeast as a model system. We show that the functional regions of yeast Are2p (the major isoform) reside in the carboxyl terminus that is most conserved in ACAT family members. We identified two conserved motifs in yeast Are2p that are essential for its enzyme activity, H/YSF and FYxDWVVN. The same two sequence motifs in human ACAT1 were also critical for activity when assayed in the are1Δ are2Δ yeast cell. The FYxDWWN motif in Are2p is likely involved in binding of fatty acyl CoA substrate since a single amino acid change in this motif caused an ∼6-fold increase in the apparent Km of the enzyme. In contrast, deletion of 30 residues from the amino terminus led to increased activity, indicating that this region is not required for activity and may play a negative, regulatory role. Antibodies against Are2p were generated. The wildtype Are2p was found to be membrane-bound, with an apparent molecular weight of about 79 kDa. No evidence of phosphorylation or N-glycosylation were found in Are2p. The HA epitope-tagged Are1 protein was also membrane-associated, and migrated as a heterogeneous protein of 55-60 kDa. The expression level of HA-Are1p was much lower than that of HA-Are2p, which correlates with its low enzyme activity. Furthermore, we demonstrate by yeast two-hybrid and co-immunoprecipitation experiments that Are1p or Are2p homo-multimerizes, but do not hetero-multimerize. We identified five candidate genes whose products may interact with Are2p in the cell.

ISBN: 9780599751682Subjects--Topical Terms:

517777
Nutrition.
Subjects--Index Terms:

ACAT

Functional characterization of sterol esterification enzymes in yeast.
LDR:03498nmm a2200397 4500 001 2323213
005 20231010062937.5
006 m o d
007 cr#unu||||||||
008 231204s2000 ||||||||||||||||| ||eng d
020 $a 9780599751682
035 $a (MiAaPQ)AAI9970202
035 $a AAI9970202
040 $a MiAaPQ $c MiAaPQ
100 1 $a Guo, Zhongmin. $3 3642209
245 1 0 $a Functional characterization of sterol esterification enzymes in yeast.
260 1 $a Ann Arbor : $b ProQuest Dissertations & Theses, $c 2000
300 $a 142 p.
500 $a Source: Dissertations Abstracts International, Volume: 62-03, Section: B.
500 $a Publisher info.: Dissertation/Thesis.
500 $a Advisor: Sturley, Stephen L.
502 $a Thesis (Ph.D.)--Columbia University, 2000.
506 $a This item must not be sold to any third party vendors.
506 $a This item must not be added to any third party search indexes.
520 $a In all eukaryotes, free sterol is an essential component of membranes. However, high levels of free sterol can be toxic. Cells maintain sterol homeostasis, in part, by converting excess free sterol to sterol ester. In mammals this reaction is mediated by the enzyme acyl-coenzyme A: cholesterol acyltransferase (ACAT). ACAT activity has been implicated in multiple physiologic processes, and in the pathological development of atherosclerosis. Two yeast ACAT homologs, ARE1 and ARE2 (for ACAT related enzymes 1 and 2, respectively), were identified, disruption of both genes produced a viable yeast cell with no detectable sterol ester. The goal of this study was to investigate the mechanism and function of sterol esterification. enzymes using yeast as a model system. We show that the functional regions of yeast Are2p (the major isoform) reside in the carboxyl terminus that is most conserved in ACAT family members. We identified two conserved motifs in yeast Are2p that are essential for its enzyme activity, H/YSF and FYxDWVVN. The same two sequence motifs in human ACAT1 were also critical for activity when assayed in the are1Δ are2Δ yeast cell. The FYxDWWN motif in Are2p is likely involved in binding of fatty acyl CoA substrate since a single amino acid change in this motif caused an ∼6-fold increase in the apparent Km of the enzyme. In contrast, deletion of 30 residues from the amino terminus led to increased activity, indicating that this region is not required for activity and may play a negative, regulatory role. Antibodies against Are2p were generated. The wildtype Are2p was found to be membrane-bound, with an apparent molecular weight of about 79 kDa. No evidence of phosphorylation or N-glycosylation were found in Are2p. The HA epitope-tagged Are1 protein was also membrane-associated, and migrated as a heterogeneous protein of 55-60 kDa. The expression level of HA-Are1p was much lower than that of HA-Are2p, which correlates with its low enzyme activity. Furthermore, we demonstrate by yeast two-hybrid and co-immunoprecipitation experiments that Are1p or Are2p homo-multimerizes, but do not hetero-multimerize. We identified five candidate genes whose products may interact with Are2p in the cell.
590 $a School code: 0054.
650 4 $a Nutrition. $3 517777
650 4 $a Cellular biology. $3 3172791
653 $a ACAT
653 $a Esterification enzymes
653 $a Sterol
653 $a Yeast
690 $a 0570
690 $a 0379
710 2 $a Columbia University. $3 571054
773 0 $t Dissertations Abstracts International $g 62-03B.
790 $a 0054
791 $a Ph.D.
792 $a 2000
793 $a English
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=9970202