Copyright is owned by the Author of the thesis. Permission is given for a copy to be downloaded by an individual for the purpose of research and private study only. The thesis may not be reproduced elsewhere without the permission of the Author.
Welcome message from author
This document is posted to help you gain knowledge. Please leave a comment to let me know what you think about it! Share it to your friends and learn new things together.
Transcript
Copyright is owned by the Author of the thesis. Permission is given for a copy to be downloaded by an individual for the purpose of research and private study only. The thesis may not be reproduced elsewhere without the permission of the Author.
ACETYL-CoA CARBOXYLASE IN PHOTOSYNTHETIC T I S SUE
A the s i s pre sented in parti al ful f i lment of the
requirements for the degree of Doctor of Philo sophy
in B iochemi stry at
MASSEY UNIVERS ITY
BASIL JOHN N IKOLAU
1 9 8 1
i i
ABSTRACT
Acetyl -CoA carboxylase catalyses the rate-limit ing reaction
in de novo f atty acid biosynthesis in a wide var iety of organisms. I n plants however , the signifi cance of this
enzyme in regulating de novo fatty ac id biosynthesis is
unknown.
In th is invest igat ion acetyl-CoA carboxylase of ma i z e , barley
and spinach leaves has been studied in order to compare some
features of this enzyme in the three pl ants . In mai ze ,
acetyl-CoA c arboxylase is located in chloroplasts , where i t occurs as a soluble enzyme in the stromal fraction . Both
mesophyll and bundle sheath ce l ls also exhibit acetyl -CoA
carboxylase activity . Activities of acetyl-CoA carboxylase
in isol ated c hloroplasts have been compared to rates of l ipid
synthesis from acetate in orde r to investigate the ro le o f acetyl-CoA carboxyl ase i n regul ating de novo fatty ac id bio
synthesis from acetate . Although acetyl -CoA carboxylase
activity was higher than that expected from the rate of
acetate incorporat ion into l ip ids of isolated chloroplasts o f mai ze and bar ley , the opposite was found for chloroplasts
isolated from spinach . In chloroplasts from the emerg ing
leaves of ma i ze seedl ings , l ipid synthesis was max imal with chloroplasts isolated from the leaf segment corresponding to
leaf greening . However , a relationship between leaf deve lop
ment and acetyl -CoA carboxylase activity was less apparent .
Consequently a regul atory role for acetyl-CoA carboxylase in de � fatty acid biosynthesis could not be d irectly estab
l ished from a compar ison of the rates of l ipid synthesis and
acetyl-CoA c arboxyl ase act iv i t ies .
An alternative approach to the establishment of a regulatory
role for acetyl -CoA carboxylase was to purify the enzyme
from mai ze and barley leaves , and study its response to var i
able concentrations o f substrates , products and certain
cellular metabolites . Pur i ficat ion of both enzymes was per-
lll
formed by identi c a l procedure s , includ ing polyethylene glycol
fract i onation , hydrophobic chromatography and ge l fi ltration .
Acetyl -CoA carboxylase from b?th mai z e and barley leave s
appeared to be an integral enzyme , a s no evidence for its
d i s sociation was found , contrary to the find i ngs of the
bacteri a l enzyme . The kinet ic properties o f the partially
puri fied enzyme from mai ze and bar ley were very similar . The
apparent Michael i s constants for the substrat e s , acetyl-CoA and HCO� , were about O . lmM and 2mM , respective ly for both
enzyme s . The enzymically active form of the substrate , ATP , was found to be Mg . ATP . Furthermore , free ATP inhibited
enzymic activity , while f ree Mg 2 + act ivated the enzyme from
both p l ant source s . Monovalent cations , particular ly K+,
were pos itive e f fectors o f acetyl-CoA carboxyl ase , on the
other hand , the products o f the acetyl -CoA c arboxylase re-
action , malonyl -CoA and ADP were inhibitor s . ADP inhibit ion
was compet itive with respect to ATP , but uncompetitive with
respect to acetyl-CoA . I nh ibition of acety l -CoA carboxylase
activ i ty by CoA was noncompet it ive with re spect to acetyl
CoA , while palmitoyl-CoA inhibition wa s uncompetit ive with
re spect to acetyl-CoA .
From the view point of regulation o f acetyl -CoA carboxylase
act iv i ty , the observed response o f activity to change s in pH ,
and the concentrat ions o f Mg2+, K
+ , ATP and ADP in vitro , may be o f s ign i f i c ance . Changes in the chloroplastic leve l s of
the se e f fecto r s of acety l-CoA carboxylase act ivity have been reported to occur during l ight-dark tran s i tion of chloro
plasts , and would be cons i stent in regulating acetyl -CoA carboxylase a c tivity in the light and dark .
iv
ACKNOWLEDGEMENTS
I wi sh to thank my superv i sor , Dr J C Hawke , for his invaluable advice , encouragement and a s s i s tance throughout the
cour se o f thi s study . I am a l so indebted to Dr C R S lack of
the Plant Physiology Divis ion , DSIR , for hi s advice and
a s s i stance , particularly dur ing the absence of Dr Hawke .
I would l ike to thank Mr I J Warr ington of the P l ant
Phys iology Divi s ion , DS I R , for helpful advice in the use o f
the c l imate control rooms for the growing o f mai ze and
barley plants .
In add it i on , particular thank s i s extended to Ms Joy Watson
for typi ng this manuscript .
ACP
ADP
AMP
ATP
BCCP
BSA
chl
P-CMB
CoA
DTT
EDTA
Hepe s
Mes
NADH
NADPH
NEM
PEG
PEP
2 -PGA
3 -PGA
LIST OF ABBREV IATIONS
acyl-carrier prote i n
adenos ine 5'-diphosphate
adenosine 5'-monophosphate
adenos ine 51-tr iphosphate
biotin carboxyl-carrier protein
bovine serum albumi n
chlorophy l l
P-chloromercuriben z oate
coenzyme A
d ithiothre itol
ethylenedi amine tetraace tic ac id
N-2-hydroxyethylpiperaz ine-N'- 2�ethane sulphonic
ac id
2[ N-Morphol i no] ethane sulphonic ac id
V
8-nicotinamide adenine dinucleotide , reduced form
n icotinamide adenine dinuc leotide pho sphate , re
duced form
N -ethylmale imide
pol�t hylene glycol
phosphoeno lpyruvate
2 -phosphoglyceric ac id
3 -phosphoglyceric ac id
PO POP
ppGpp
PPO
pppGpp
RNA
RuDP
TEMED
TLC
Tr i c in e
Tr i s
1 , 4 -bi s[2 ( 5 -phenylox a z o l y l )]Benzene
guano s ine 5 ' -d ipho sphate- 3 ' -d ipho sphate
2 , 5 -diphenyloxazole
guanos ine 5 ' -triphosphat e- 3 ' -d ipho sphate
r ibonucleic ac id
ribu lose 1 , 5 -diphosphat e
N , N , N ' , N'-Tetramethylethy lened iamine
thin-layer chromatography
N -tris[Hydroxymethyl]-methyl glyc ine
t r i s ( hydroxymethyl ) aminomethane
vi
vii
TABLE OF CONTENTS
AB STRACT i i
ACKNOWLEDGEMENTS iv
L I ST OF ABBREVIAT IONS
TABLE OF CONTENTS
L I ST OF F I GURES
LIST OF TABLES
CHAPTER 1 INTRODUCT ION
1 . 1 General Background
V
Vll
x i i i
xvi i i
1
1 . 2 Acety l -CoA Carboxylase from An imal Sources
1 . 2 . 1 Mo lecu lar prope rties 2
1 . 2 . 2 Regu lat ion o f cata lytic activity
1 . 2 . 2 . 1 Long-term control o f
acetyl-CoA carboxylase 2
1 . 2 . 2 . 2 Short-term control of
acetyl -CoA carboxy lase 4
1 . 3 Acetyl -CoA Carboxylase from Escherichia col i
1 . 3 . 1 Molecular propert ies 8
1 . 3 . 2 Regulat i on o f cata lytic act iv ity 9
1 . 4 Acetyl -CoA Carboxylase from Yea st 10
1 . 5 Acet y l-CoA Carboxyl ase f rom Plant s 1 1
1 . 6 Acetyl -CoA Carboxylase from Other S ources 1 2 .
1 . 7 Acety l -CoA Carboxylase a s a B iot in Enzyme 1 4
1 . 8 Fatty Ac id Biosynthe s i s in Plants 1 5
CHAPTER 2 MATERIALS
2 . 1 P lant Mater i a l s 2 0
CHAPTER
viii
2.2 Reage nts
3 METHODS
3.1 Preparation o f Acy l -CoA Esters
3.2 En z yme Assays
3. 3 Preparation o f C e l l - free Extracts
3. 4 Preparation o f Chloropl asts
3.5 Preparation o f Me sophyl l Protop lasts and
Bund le Sheath Strand s
3. 6 Preparation o f E . co l i Acety l-CoA Carbox
lase Component s
3.7 I ncorporation o f [ l - 1 �C ] Acetate into Lipids
3. 7. 1 [ l - 1 4C ] acetate incorporat ion into
2 0
2 2
2 3
2 5
2 6
2 6
2 8
lipids by i solated chloroplasts 2 9
3. 7. 2 [ l- 1 4C] acetate incorporat i on into
l ipids by leaf s l i ces
3. 8 Preparation of Ma i z e Lea f Section s
3. 9 Preparation o f S epharo se -N-Propane
3.1 0 Polyacry lamide Ge l Electrophere s i s
3.1 1 Centr ifugat ion
3.1 2 Ana l yti cal Methods
3.1 2. 1 Determinat i on o f prote in
3.1 2. 2 Determinat i on of chlorophy l l
3.1 3 Determination o f Radi oact ivity
3.1 4 Chromatographic P rocedure s
3.1 4. 1 Gel f i ltrat ion chromatography
3.1 4. 2 Thin�layer chromatography
3.1 4. 3 P aper chromatography
3.1 5 Pur i f i cation o f Acetyl-CoA Carboxylase
from Leave s
2 9
3 0
3 0
3 0
3 1
3 2
3 2
3 3
3 3
3 3
3 3
3 4
CHAPTER 4 RESULTS
GENERAL FEATURE S AND STAB I LITY OF LEAF
ACETYL-CoA CARBOXYLASE
4.1 General Feature s of Acetyl -CoA Carboxy
lase Act iv ity
4.2 Stab i l i zat ion o f Acetyl -CoA Carboxylase
in Ce l l -Free Extract s by Sul fhydryl Pro
tect ing Agent s
LOCAT I ON OF ACETYL-CoA CARBOXYLASE ACTIVITY
IN MAI Z E LEAVES
4 . 3 Locat ion of Acety l -CoA Carboxy lase in
Chlorop last s o f Ma i z e Leave s
4. 4 Solub i l ity o f Ma i z e Chloropl astic Acetyl
CoA Carboxylase
4 . 5 Location o f the B iot in Prosthetic Group
ix
3 6
4 2
4 6
4 8
o f Chloroplast i c Acetyl -CoA Carboxylase 5 1
4. 6 Distr ibut ion o f Ac ety l -CoA Carboxyl a s e
Between Mesophy l l and Bund le She at h C e l l s
o f Mai z e Leave s
LEVELS OF ACETYL-CoA CARBOXYLASE ACTIVITY IN
LEAVES
4 . 7 I nve st igat ion o f the Re lat ionship B etween
Leaf Deve lopment , Acetyl -CoA Carboxylase
Activity and [ l - 1 4C] acetate Incorpor ation
5 5
into Lipids 5 5
4. 7.1 I ncorporat ion o f [ l- 1 ljC] acetate
by ma i z e l e a f s l ic e s 6 0
4. 7. 2 I ncorporat i on o f [ l - 14 C J acetate
into l ip i d s by i solated chloro
p l a sts from deve loping ma ize
l eave s
4.7.3 Acetyl -CoA carboxylase activity
in c e l l - free extract s of deve lop
ing mai ze l eaves
6 0
6 2
4 . 7 . 4 Acety l -CoA carboxylase act ivity in
isolated chloropla st s from deve lop
ing ma i z e leave s
4 . 8 Acetyl-CoA Carboxyla se Activity and Lipid
Synthe s i s in Leave s o f Ma i z e , Barley and
X
6 5
Spinach 6 5
4 . 9 E f fect o f Light on Ac etyl-CoA Carboxylase
Act ivity in Ma i z e Leave s
PURIFICATI ON OF LEAF ACETYL-CoA CARBOXYLASE
4 . 1 0 Pur i f ic at ion o f Acety l -CoA Carboxy l a s e
from Ma i z e Leave s
4 . 1 1 Pur i f i c at ion o f Ac etyl -CoA Carboxy lase
from Barley Leave s
K I NET IC PROPERT IES OF ACETYL-CoA CARBOXYLASE
4 . 1 2 E f fect of Acety l -CoA and HC03 Concentra-
7 0
7 0
8 3
t ions on Acetyl -CoA Carboxylase Act ivity 8 4
4 . 1 3 E f f ect o f ATP and MgC12 Concentrat ions
on Acety l -CoA Ca rboxylase Act iv ity
4 . 1 4 Spec i f i c ity of the Divalent Metal Ion
Requi red for Acety l-CoA Carboxylase
Act ivity
4 . 1 5 E f fect of pH on Acetyl -CoA Carboxylase
Act ivity
4 . 1 6 E f fect of Temperature on Acetyl -CoA
Carboxyl a se Act ivity
4 . 1 7 Act ivation of Acetyl -CoA Carboxylase
Activity by Monova l ent Cat ions
4 . 1 8 S ubstrate Spec i f i c ity of Acetyl-CoA
Carboxy l a s e
4 . 1 9 I nhibition o f Ma i z e L e a f Acetyl - CoA
Carboxylase by Ma lonyl -CoA
9 2
1 0 5
1 0 5
1 0 7
1 0 7
1 1 1
1 1 1
4 . 2 0 Effect o f ADP on Acety l -CoA Carboxy lase
Act ivity
4 . 2 1 E f fect of CoA on Acetyl -CoA Carboxyl a s e
Act ivity
4 . 2 2 E f f ect of Palrnitoyl -CoA on Acetyl -CoA
Carboxy l a s e Act ivity
4 . 2 3 E ffect of AMP , Pho sphoenolpyruvate ,
Oxaloacet ate and Citrate on Acetyl -CoA
Carboxyla s e Act ivity
4 . 2 4 E f f ect o f Sul fhydryl Reacting Reagent s
on Acetyl -CoA Carboxylase Act ivity
CHAPTER 5 DI SCUS S I ON
5 . 1 Acetyl-CoA Carboxylase - it s General
Feature s a nd Stabi l ity
5 . 2 Cellular Location o f Acetyl-CoA Carboxy
lase in Ma i ze Le ave s
5 . 3 Re l ationship Between Acetyl-CoA Carboxy
l ase Act iv ity and Lipid Synthe s i s in
I so l ated Ch loropl a st s
5 . 4 Re lation sh i p Between Acetyl-CoA Carboxy
lase Act ivity , Lipid Synthe s i s and Lea f
Deve lopment
x i
1 1 6
1 1 6
1 2 5
1 2 5
1 2 8
1 3 0
1 3 2
1 3 5
1 3 7
5 . 5 Pur i f icat ion o f Acety l -CoA Carboxylase 1 3 9
5 . 6 K inetic Properties o f Acetyl-CoA Carboxy-
lase
5 . 6 . 1 E f f e ct of sub strate s on acety l -CoA
carboxylase 1 4 2
5� 6 . 2 Sub strate spec i f icity o f acetyl -
CoA c arboxy lase
5 . 6 . 3 E f fectors o f acety l-CoA c arboxy
l a s e a ctivity
5 . 7 L i ght-Dark Modulation o f Acetyl - CoA
Carboxy l a s e Act ivity
1 4 3
1 4 4
1 4 6
5 . 8 Summary and Sugge st ions for Further
Study
BIBLIOGRAPHY
x i i
1 4 8
1 5 1
x i i i
LI ST O F FIGURE S
F igure Page
1 Structura l c l a s s i f ication of biot in c ontaining
e n zyme s
2 I dent i f icat i on o f the product of acetyl -CoA
c arboxy l a s e as say by TLC
3 Identificat ion o f the product of the acety l -
1 6
3 7
CoA carbox y l ase a s say by paper chromatography 3 8
4 Dependence o f acetyl -CoA c arboxylase activity
on protein l eve l s of cell- free extracts o f
mai ze leave s
5 E f fect o f t ime on acetyl-CoA carboxyl a s e
4 0
act ivity i n c e l l - free extracts of ma i z e leaves 4 1
6 Stab i l i z at ion o f acetyl -CoA carboxylase act iv
ity in ce l l - free extracts of ma ize by 8 -mer-
c aptoethano l 4 3
7 Stabi l i zat i on o f acetyl -CoA carboxylase act iv
ity in cel l - free extracts of barley by 8 -mer
c aptoethano l
8 D i stribut ion o f enzymes , chlorophy l l and pro
te in fol lowing the centrifugat ion of pur i f ied
ma i ze chloropl ast s in a suc ro s e density
gradient
9 Repre sentat ive f i e ld o f a preparation o f i so
l ated mai z e chloroplasts
4 4
4 7
4 9
x iv
Figure Page
1 0 D i str ibut ion o f enzyme s , chlorophy l l and
prote in f o l lowing the centri fugation of d i s
rupted ma i z e chlorop l asts i n a di scont i nuou s
sucro se den s ity gradient 5 0
1 1 Representat ive f i e ld o f a preparation o f iso
l ated mai z e me sophyl l protopl ast s
1 2 Representat ive f i e ld o f a preparation o f mai z e
5 8
bundle sheath strands 5 9
1 3 E f fect o f l ight on the activity o f acety l -CoA
c arboxyla s e in ma i z e leave s
1 4 E lut ion o f a c etyl -CoA carboxylase of ma i z e from
S epharo se-N -Propane
1 5 Ge l f i ltrat ion o f acety l -CoA carboxy l a s e o f
ma i z e on U lt rogel AcA2 2
1 6 Ge l f i ltrat i on o f acetyl -CoA carboxy l a s e o f
mai z e on U ltrogel AcA2 2
1 7 Polyacrylamide gel e l ectropher e s i s o f acetyl
CoA c arboxyl as e o f ma i z e l eave s
1 8 Rel at ionship b etween HC03 , acetyl-CoA concen
trat i ons and the activity o f ma i ze leaf
acet y l-CoA c arboxylase
1 9 Re l at ionship between HC03 , acety l-CoA concen
trat ions and the activity o f bar l ey leaf
acetyl -CoA carboxylase
2 0 Lineweaver -Burk analy s i s o f the e f fect o f
acetyl -CoA on the activity o f maize leaf
acet y l-CoA c arboxy lase
7 1
7 7
7 8
8 0
8 1
8 6
8 7
8 8
XV
Figure Page
2 1 Lineweaver -Burk analy s i s o f the e f fect o f
acety l -CoA o n t h e act ivity o f bar ley l e a f
acety l - CoA carboxy lase
2 2 Re l at ionshi p between HC03 , acety l -CoA concen
trat ions and the activity o f ma i z e leaf
acety l - CoA carboxyl ase
2 3 Re lat ionship be tween HC03 , ac etyl -CoA conc en
trat ions and the activity of bar ley l e a f
acetyl -CoA c arboxylase
2 4 Lineweaver -Burk analy s i s o f the e f fect o f
HCO� o n the activity o f ma i z e leaf acetvl-CoA �
carbox y l a s e
2 5 Lineweaver -Burk a naly s i s o f the e f fect o f
HCO� on the activity of bar ley leaf acetyl -CoA
carboxylase
2 6 Effect o f MgC 1 2 o n the act ivity o f ma i z e leaf
acety l - CoA carboxylase
2 7 Effect o f MgC l 2 o n the act ivity o f barley l e a f
acetyl -CoA carboxylase
2 8 Ef fect o f ATP o n the act ivity o f ma i z e leaf
acety l - CoA c arboxylase
2 9 E f fect o f ATP on the activity o f bar ley leaf
acety l-CoA carboxy lase
3 0 Re lationship between ATP , MgC l 2 concentration s
and t h e activity o f mai z e l e a f acety l - CoA
carbox y l a s e
8 9
9 0
91
9 3
9 4
9 5
9 6
9 7
9 8
1 0 0
xvi
Figure Page
3 1 Re l ation ship between ATP , MgC l2 concentrations
and the act ivity of barley leaf acety l -CoA
carboxy l a s e
3 2 Ana ly s i s b y Hil l -plot of the relation s h ip be
tween ATP , MgC12 concentrat ions and the
activity of ma ize l e a f acetyl -CoA carboxylase
3 3 Ana ly s i s by Hill-plot o f the re lat ions h ip be
tween ATP , MgC l2 concentrat ions and the activ-
1 0 1
1 0 2 , 1 0 3
i ty o f bar l ey leaf acetyl-CoA carboxyl a s e 1 0 4
3 4 E f fect o f d iva lent metal ions o n the a c t ivity
o f ma i z e l ea f acety l -CoA carboxylase
3 5 E ffect o f pH on the act ivity o f ma i z e l e a f
acety l -CoA c arboxy l a se
3 6 E f fect o f pH on the act ivity o f bar l ey l e a f
acetyl -CoA carboxyl a s e
3 7 E f fect o f temperature on the activity o f ma i z e
1 0 6
1 0 8
1 0 9
l e a f acetyl -CoA c arboxylase 1 1 0
3 8 E f fect o f KCl on the activity o f ma i z e l e a f
acety l -CoA carboxylase
3 9 Spec ific ity of carboxylation by ma i z e l e a f
a c e tyl -CoA carboxylase
4 0 E f fect o f malonyl-CoA on the activity o f mai z e
l e a f acetyl -CoA carboxylase
4 1 ADP inhibition o f the ac tivity o f ma i z e l e a f
acetyl-CoA carboxylase
1 1 2
1 1 4
1 1 5
1 1 7
xv ii
F igure Page
4 2 L i neweaver-Burk analys i s o f the e f fect o f ADP
on the activity o f ma i z e leaf acety l-CoA
c arboxylase
4 3 Ana l y s i s by Dixon-plot o f the e f fect o f ADP
�n the act ivity o f ma i z e leaf acety l - CoA
c arboxylase
4 4 ADP inhibi t ion o f the a ctivity o f ma i z e l e a f
acety l -CoA c arboxyl a s e
4 5 L ineweaver-Burk ana l y s i s of the e f fect o f ADP
o n the act ivity o f ma i z e leaf acetyl-CoA
c arboxylase
4 6 CoA inhibition o f the activity o f ma i z e leaf
acetyl -CoA carboxyl ase
4 7 L ineweaver-Burk ana ly s i s of the ef fect o f CoA
o n the act ivity of ma i z e leaf a cety l - CoA car-
1 1 8
1 1 9
1 2 0
1 2 1
1 2 2
boxylase 1 2 3
4 8 Ana l y s i s by D ixon-plot o f the e f fect o f CoA
o n the activ ity o f ma i z e leaf acety l - CoA car-
boxy l ase 1 2 4
4 9 P a lmitoyl -CoA inhibition of the activity o f
ma i z e leaf acetyl-CoA c arboxylase
5 0 L i neweaver -Burk analys i s of the e f f ec t o f
pa lmitoyl-CoA on the activity o f ma i z e leaf
acetyl-CoA c arboxylase
1 2 6
1 2 7
xv i ii
Table
I
L I ST OF TABLES
Protomer-polyrner tran s i t ion of an imal
acetyl -CoA carboxylase
I I Co f actor requ irements o f acetyl-CoA c arboxy-
5
l a s e activity in cel l - free extracts o f ma i z e 3 9
I l l Ace ty l -CoA c arboxylase ac t ivity i n c e l l - free
extracts o f sp inach l eave s
IV
V
Preparation o f biot in c arboxylase from E . coli
Co f ac tor requ i rements for the carboxylation of
the biotin pro s thetic group in ma i z e chloro
p l a s t s by biot i n c arboxylase of E. col i
VI Ma l onyl -CoA format ion from carboxyl -biotin by
mai z e chloropl a st s
VI I D i s t r ibut ion o f the biotin pro sthet ic group in
chloroplasts i so l ated from the leave s o f three
4 5
5 2
5 3
5 4
p l an t s 5 6
VI I I D i s tribution o f en zymes between me sophy l l proto-
p l a s t s and bundle sheath s trand s 5 7
IX E f fe c t of leaf deve lopment on [l - 1 4 C ] acetate
incorporation into l ip i d s by leaf s l i c e s of
ma i z e ·
X E f fect of l ea f deve lopment on [l - 1 4C ] acetate
incorporat ion i nto l ipids by i sol ated chloro
p l a s t s of mai z e
6 1
6 3
Tabl e
XI Acety l-CoA carboxylase act ivity in c e l l - free
extracts obta ined from section s of maize
leave s
XI I Acetyl -CoA carboxylase act ivity in c hloro-
xix
6 4
pl ast s isolated from sect i ons o f ma i ze leave s 6 6
XI I I [l- 1'"'C J acetate incorporat ion into l ipids o f
chloropl asts and acetyl -CoA carboxyla se
act iv ities in c e l l - free extracts and chloro
pl ast s from ma i z e , bar l ey and spinach
XIV Acety l-CoA carboxylase in ma i z e and spinach
leave s
XV Pur i f ication o f acetyl-CoA carboxylase of
ma i z e leave s us ing procedure I
XVI Pur i f ication o f acety l-CoA carboxy lase of ma i z e
leave s us ing proc edure I I
XVI I
XVI I I
I nte ract ion of ac etyl-CoA c arboxyl ase with
Sepharo se-N-Cx
Stabil ity of part ially pur i f ied ac etyl -CoA
carboxylase
XIX Pur i f ication o f acetyl-CoA carboxyl a s e of
barley leave s u s i ng procedure I
XX Act ivation of ma i z e lea f acetyl-CoA c arboxy
lase by monovalent cat ions
XXI Inhibition of ma i z e leaf ac etyl -CoA c arboxy-
6 7
6 9
7 3
7 4
7 5
8 2
8 5
1 1 3
lase act ivity b y sulfhydryl reacting reagent s 1 2 9
Related Documents
