A Defect in Branched-Chain Amino Acid
Metabolism in a Patient with Congenital Lactic
Acidosis due to Dihydrolipoyl Dehydrogenase
Deficiency
JENNIFER TAYLOR, BRIAN H. ROBINSON,"5' AND W. GEOFFREY SHERWOOD
Departments of Paediatrics and Biochemistry, University of Toronto; and Research Institute, The Hospital for Sick
Children, Toronto, Ontario, Canada
Summary
In a case of dihydrolipoyl dehydrogenase deficiency, there
was not only an elevation of lactate and a-ketoglutarate but
also of branched chain amino acids. The levels of branchedchain amino acids varied from the normal range to three times
the upper limit of normal during the patient's lifetime, and alloisoleucine was detectable at all times. Examination of postmortem tissues revealed that the activity of branched-chain keto
acid dehydrogenases was between zero and 10% of that in
control tissues. It is suggested that the multiple defects seen in
oxidative decarboxylation in this patient is the consequence of a
single genetic deletion of an enzyme common to pyruvate
dehydrogenase, a-ketoglutarate dehydrogenase, and branchedchain keto acid dehydrogenases.
Speculation
The dihydrolipoyl dehydrogenase component of pyruvate, aketoglutarate, and branched-chain keto acid dehydrogenases is
genetically and biochemically a single entity.
o f a-keto-acids have a s i m i l a r f u n c t i o n a l make-up c o n s i s t i n g i n
each c a s e of t h r e e main c a t a l y t i c enzymes bound t o g e t h e r i n a
d i s c r e t e multienzyme complex.
The f i r s t of t h e s e enzymes,or
a-keto-acid d e c a r b o x y l a s e ( E l ) , r e q u i r e s t h i a m i n e pyrophosphate f o r
a c t i v i t y and i s r e s p o n s i b l e f o r t h e d e c a r b o x y l a t i o n o f t h e
c a r b o x v l f u n c t i o n a d i a c e n t t o t h e k e t o n e aroun.
The second enzvme
( E 2 ) . 6r d l h y d r o l r p o ; l t r a n s a c e t y l a s e , ha: boAund l l p o a t e r e s l d u e s
and is responsible f o r t r a n s f e r r i n g t h e d e c a r b o x y l a t e d f u n c t i o n t o
a coenzyme A moiety.
The t h i r d enzyme ( E 3 ) , o r d i h y d r o l i p o y l
dehydrogenase, i s r e s p o n s i b l e f o r t r a n s f e r of r e d u c i n g hydrogen
El i s l i k e l y
from l i p o a t e t o a f l a v o p r o t e i n and u l t i m a t e l y NAD.
t o be a d i f f e r e n t enzyme i n each of t h e t h r e e d i f f e r e n t k e t o a c i d
dehydrogenase complexes s i n c e it d e a l s w i t h s t r u c t u r a l l y d i f f e r e n t
k e t o a c i d s u b s t r a t e s , and i n inborn e r r o r s o f p y r u v a t e decarboxyl a s e (1.4) and branched-chain k e t o a c i d d e c a r b o x y l a s e ( 9 ) a r e
s e p a r a t e e n t i t i e s . E2 is a l s o l i k e l y t o be a d i f f e r e n t enzyme i n
each of t h r e e complexes, a g a i n because it h a s t o d e a l w i t h
structurally different substrates.
Eg a c t i v i t y , however, i n v o l v e s
t h e same s u b s t r a t e i n a l l t h r e e complexes, it i s d e f i c i e n t i n o u r
p a t i e n t , and i t s d e f i c i e n c y e x p l a i n s a l l of t h e enzymic d e l e t i o n s
we have demonstrated i n t h i s p a t i e n t .
We r e c e n t l y r e p o r t e d a c a s e of c o n g e n i t a l l a c t i c a c i d o s i s
which was found t o be due t o a d e f i c i e n c y of t h e t h i r d component
of t h e p y r u v a t e dehydrogenase complex ( E j ) . T h i s d e f i c i e n c y was
found t o compromise t h e a c t i v i t y a l s o of t h e a - k e t o g l u t a r a t e
dehydrogenase complex: i n d i c a t i n g t h a t d i h y d r o l i p o y l dehydrogenase
i s s h a r e d i n common by t h e s e two enzymes ( 8 ) . The p a t i e n t
concerned had e l e v a t e d l e v e l s o f branched-chain amino a c i d s
i n d i c a t i n g a p o s s i b l e involvement of t h e branched-chain k e t o a c i d
d e h v d r o ~ e n a s e s i n t h e ~ a t h o s e n e s i sof t h i s d i s e a s e .
In t h i s b r i e f
c o & u n i c a t i o n we demonstrat; i n p o s t mortem t i s s u e from t h i s
p a t i e n t t h a t t h e branched-chain k e t o a c i d dehydrogenase a c t i v i t y
1s d e f i c i e n t and t h a t t h e d e f e c t i s n o t a s s o c i a t e d w i t h t h e d e c a r b o x y l a s e p o r t i o n o f t h e enzyme complex.
CASE REPORT
D e t a i l s of t h e c a s e r e p o r t a r e d e s c r i b e d i n r e f e r e n c e ( 8 ) .
Levels of branched c h a i n amino a c i d s throughout t h e l i f e o f t h e
p a t i e n t a r e shown i n F i g u r e 1.
MATERIALS AND METHODS
REFERENCES
Branched-chain k e t o a c i d dehydrogenase was measured by t h e
modified method of T a y l o r e t a l . ( l l ) , u s i n g (1-14~1-branchedc h a i n k e t o a c i d s p r e p a r e d from t h e corresponding l - 1 4 ~ - a m i n o a c i d s
by t h e method o f Rudiger e t a l . ( 9 ) . Branched-chain k e t o a c i d
d e c a r b o x y l a s e was measured by t h e modified method of Reed and
W i l l m s (7).
RESULTS
PLASMA AMINO ACID LEVELS
The l e v e l s o f a l l t h r e e branched-chain amino a c i d s were
measured s e v e r a l t i m e s d u r i n g t h e p a t i e n t ' s f i v e months i n h o s p i They appeared t o be uniformely e l e v a t e d on f i v e o u t o f t h e
tal
e i g h t t i m e s t h a t t h e y were measured ( F i g . 1 ) ; on two o c c a s i o n s t o
markedly h i g h l e v e l s .
Levels o f a l l o i s o l e u c i n e of 1 0 , 14 and 14uM
were d e t e c t e d on t h r e e o c c a s i o n s , t h e r e b e i n g a t r a c e a t t h e o t h e r
t i m e s of measurement.
Alloisoleucine is undetectable i n t h e
plasma o f t h e normal i n d i v i d u a l .
.
POST MORTEM STUDIES
B l a s s , J.P., Avigan, J., and Uhlendorf, B.W.: A d e f e c t i v e
p y r u v a t e d e c a r b o x y l a s e i n a c h i l d w i t h an i n t e r m i t t e n t movement d i s o r d e r . J. C l i n . I n v e s t . 49:423, 1970.
Dancis, J., H u t z l e r , J . , and L e v i t z , M.:
Thin layer-chromatography and s p e c t r o p h o t o m e t r y of a - k e t o - a c i d hydrazones.
Biochim. Biophys. Acta 78:85, 1963.
Dancis, J . , and L e v i t z , M.: A b n o r m a l i t i e s o f branched-chain
amino a c i d metabolism.
I n : S t a n b u r y , J.B., Wyngaarden, J . B . ,
and F r e d r i c k s o n , D.S. ( e d s . ) , The M e t a b o l i c B a s i s of I n h e r i t e d
D i s e a s e , McGraw-Hill, New York, p. 426 ( 1 9 7 2 ) .
F a r r e l , D.F., C l a r k , A.F., S c o t t , C.R., and Wennberg, R.P.:
Absent p y r u v a t e d e c a r b o x y l a s e i n man.
A cause of congenital
l a c t i c acidosis.
S c i e n c e 187:1082, 1975.
Khatra, B.S., Chawla, R.K.,
S e w e l l , C.W., and Rudman, D.:
D i s t r i b u t i o n of branched-chain a-keto-acid dehydrogenase i n
J. C l i n . I n v e s t . 59:558, 1977.
primate t i s s u e s .
L a n c a s t e r , G., Mamer, O.A., and S c r i v e r , C.R.:
Branched-chain
a - k e t o - a c i d s i s o l a t e d a s oxime d e r i v a t i v e s .
Relationship t o
t h e c o r r e s p o n d i n g hydroxy a c i d s and amino a c i d s i n Maple Syrup
Urine D i s e a s e . Metabolism 23:257. 1974.
Reed, L.J., and W i l l m s , C.R.: Pyruvate d e c a r b o x y l a s e .
Methods.
Enzymol. 9:258, 1966.
Deficiencv of
Robinson. B.H.. T a v l o r . J . . and Sherwood. W.G.:
d i h y d r o l i p o y l dehyhroqenaa; ( a component.of t h e p y r u v a t e a6d
a - k e t o g l u t a r a t e dehydrogenase complexes): A c a u s e o f congenit a l chronic l a c t i c - a c i d o s i s i n infancy.
P e d i a t . Res. (1977,
i~ p r e s s ) .
Rudiger, H.W.,
Langenbeck, U . , Schulze-Schencking, M . ,
Goedde, H.W., and Schuchmann, L.:
Defective decarboxylase i n
branched-chain k e t o - a c i d o x i d a s e multi-enzyme complex i n
c l a s s i c a l t y p e s o f Maple Syrup Urine D i s e a s e .
Humangenetic
14:257, 1972.
Schulman, J . D . ,
L u s t b e r g , T.J., Kennedy, J . L . , Museles, M . ,
and S e e q m i l l e r , J.E.:
A new v a r i a n t o f Maple Syrup Urine
D i s e a s e (branched-chain k e t o a c i d u r i a ) .
Amer. J. Med. 49:118,
1970.
T a y l o r , S . I . , Mukherjee, C., and J u n g a s , R.L.:
S t u d i e s on t h e
mechanism of a c t i v a t i o n of a d i p o s e t i s s u e p y r u v a t e dehydrogenJ. B i o l . Chem. 248:73, 1973.
a s e by i n s u l i n .
Van den H u r s t , J.L. and Wadman, S.K. : A v a r i a n t form o f
branched-chain k e t o a c i d u r i a .
Acta Paed. Scand. 60:594. 1971.
W e s t a l l , R.G., Dancis, J . , and M i l l e r , S.:
Maple Suga; Urine
D i s e a s e . Amer. J . Dis. C h i l d . 94:571, 1957.
We thank t h e Canadian Medical Research C o u n c i l and t h e Weston
Foundation f o r f i n a n c i a l s u p p o r t , and t h e Department o f
C l i n i c a l B i o c h e m i s t r y , The H o s p i t a l f o r S i c k C h i l d r e n , i n
p r o v i d i n g f a c i l i t i e s f o r amino a c i d a n a l y s i s .
.
T o t a l a - k e t o i s o c a p r o a t e dehydrogenase measured on p o s t mortem
kidney, l i v e r , and b r a i n showed t h a t t h e p a t i e n t had l e s s t h a n 10%
o f normal a c t i v i t y i n each c a s e compared t o t h e o t h e r c o n t r o l s ,
Table 1. a - K e t o i s o c a p r o a t e d e c a r b o x y l a s e . on t h e o t h e r hand.
appeared t o be i n t h e normal range i n b o t h b r a i n and k i d n e y , b u t
appeared t o b e d e p r e s s e d i n t h e l i v e r .
The a c t i v i t i e s of a-ketoB-methylvalerate dehydrogenase and a - k e t o i s o v a l e r a t e dehydrogenase
were a l s o l e s s t h a n 10% of normal a c t i v i t y i n l i v e r and b r a i n
( d a t a n o t shown). Enzyme a c t i v i t i e s o b t a i n e d f o r branchLChain
k e t o a c i d dehydrogenase i n t h e c o n t r o l t i s s u e s were s i m i l a r t o
2. ( 5 ) f o r human t i s s u e s .
t h o s e r e p o r t e d by Khatra
9.
10.
DISCUSSION
The p a t i e n t s u f f e r i n g from a d e f i c i e n c y of d i h y d r o l i p o y l
dehydrogenase ( E 3 ) , d e s c r i b e d i n r e f e r e n c e (81, a p p e a r s a l s o t o
have a d e f e c t i n branched-chain amino a c i d metaboiism.
By
measurement o f t h e branched-chain k e t o a c i d dehydrogenases, we
have shown t h a t t h i s p a t i e n t h a s l e s s t h a n 10%o f t h e normal
a c t i v i t y e x h i b i t e d by t h e s e enzymes i n l i v e r , b r a i n , and kidney.
S e v e r e d e f i c i e n c y ( < 2 %normal) a t t h i s p o i n t i n t h e metabolism of
branched c h a i n amino a c i d s l e a d s t o c l a s s i c a l Maple Syrup Urine
D i s e a s e (MSUD) ( 2 , 9 , 1 3 ) , w h i l e l e s s s e v e r e d e f i c i e n c y (2-8%) g i v e s
a v a r i a n t MSUD w i t h low p r o t e i n t o l e r a n c e , and d e f i c i e n c y a t 8-15%
g i v e s a v a r i a n t MSUD w i t h normal p r o t e i n t o l e r a n c e ( 3 , 1 0 , 1 2 ) .
Though t h i s p a t i e n t had e l e v a t e d plasma branched-chain amino a c i d s
t h e l e v e l s were a t no time g r e a t e r t h a n 0.8 mM, which i s i n d i c a t i v e of a mild form o f MSUD ( 6 ) . As f a r a s we can a s s e s s t h i s was
n o t a major f a c t o r i n t h e d i s e a s e p r o c e s s which l e d t o h i s demise.
11.
12.
13.
14.
15. Reauests f o r r e n r i n t s s h o u l d be a d d r e s s e d t o :
B r i a n H.
~ o g i n s o n ,P ~ . D . ; Research I n s t i t u t e , The H o s p i t a l - f o r S i c k
C h i l d r e n , 555 U n i v e r s i t y Avenue, T o r o n t o , O n t a r i o , M5G 1x8.
Canada.
INTERPRETATION OF MULTIPLE DEFECTS I N OXIDATIVE DECARBOXYLATION
I t h a s now been shown by us t h a t t h i s p a t i e n t was compromised a t t h r e e i m p o r t a n t s i t e s , p y r u v a t e dehydrogenase.
a - k e t o g l u t a r a t e dehydrogenase ( 8 ) . and branched-chain k e t o a c i d
dehydrogenase. A l l of t h e s e enzymes f o r o x i d a t i v e d e c a r b o x y l a t i o n
16.
Received f o r p u b l i c a t i o n June 8.1977.
17.
Accepted for p u b l i c a t i o n September 22,1977.
TABLE 1
a-KETOISOCAPROATE DEHYDROGENASE (DH) , a-KETOISOCAPROATE DECARBOXYLASE (DC)
AND DIHYDROLIPOYL DEHYDROGENASE (E3) I N POST MORTEM TISSUE
KIDNEY
LIVER
BRAIN
nmol/min/gm t i s s u e
DH
DC
3
DH
DC
E3
DH
DC
0.19
260
0.67
0.42
E3
Patient
0.69
0.16
117
Controls 1
4.9
0.16
2620
19.4
0.53
6120
5.9
0.51
3750
2
14.2
0.46
10800
19.6
0.90
14490
21.9
0.55
3830
0.80
80
3
9.1
0.25
9050
13.4
0.46
13200
5.8
0.47
2480
4
3.6
0.10
5850
12.2
0.58
7978
7.3
0.37
2570
5
3.9
0.07
4.9
0.28
-
-
-
Eg v a l u e s a r e t r a n s p o s e d from r e f e r e n c e ( 8 ) .
ISOLEUCINE
months
Figure 1.
months
months
Levels of branched-chain amino a c i d s i n t h e plasma of
t h e p a t i e n t from t h e second month of l i f e through t o
t h e seventh.
The hatched b a r s i n d i c a t e t h e range of
normal v a l u e s f o r t h e second t o t h e f o u r t h month,
normal values f o r t h e f o u r t h t o t h e seventh month a r e
not available.
Copyright O 1978 International Pediatric Research Foundation, Inc.
Printed in U.S.A