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Acid hydrolases in serum from patients with lysosomal disorders
33
Clinica Chimica Actu, 100 (1980) 33-38 0 Elsevier/North-Holland Biomedical Press
CCA 1196
ACID HYDROLASES DISORDERS
IN SERUM FROM PATIENTS
WITH LYSOSOMAL
B. HULTBERG aq*, A. I~AKS~~N b, s. SJ~~BLAD e and P.A. ~~CKERMAN a Departments of Clinical Chemistry a and Pediatrics c, University Hospital, S-221 85 Lund (Sweden), and Department of Clinical Chemistry b, Central Hospital, S-351 85 Vtixjii (Sweden)
(Received July 3rd, 1979)
summary The activity of acid hydrolases was studied in serum from patients with mucolipidosis (II and III) and other lysosomal disorders. In mucolipidosis II and III all hydrolases examined except a-glucosidase, fl-glucosidase and acid phosphatase were greatly increased. High values for P-galactosidase were seen in mucopolysaccharidosis types I and II, Gaucher’s disease, juvenile amaurotic idiocy and metachromatic leucodystrophy. N-Acetyl-fl-glucosaminidase activity was high in mucopolysaccharidosis types I, II, III and Gaucher’s disease. The activity of /I-glucuronidase was increased in mucopolysaccharidosis types I, II and III, Gaucher’s disease, juvenile amaurotic idiocy and metachromatic leucodystrophy. Acid phosphatase had increased activity only in Gaucher’s disease. In several lysosomal storage disorders no increased values could be found. It is suggested that high values in serum from patients with lysosomal storage disorders (not including mucolipidosis II and III) may depend upon liver cell damage, which disturbs the clearing of acid hydrolases from serum.
Introduction I-cell disease (mucolipidosis II) and mucolipidosis III are especially interesting lysosomal disorders because they provide a model for the exploration of basic endo- and exocytosis phenomena of acid hydrolases. One of the main features of mucolipidosis II and III is that lysosomal enzymes are elevated in
* Correspondence Sweden.
to: Dr. BjBm Hultberg,
Dept. of Clinical Chemistry,
University
Hospital,
S-221
85 Lund.
34
the medium of cultured fibroblasts from patients with the disease and are also greatly elevated in the serum of these patients [ 1,2]. A slight increase in the activity of acid hydrolases in serum has also been found in some other lysosomal storage disorders, in patients with Gaucher’s disease [3], mucopolysaccharidosis [4,5], and in sera from patients with juvenile amaurotic idiocy [6]. The comparatively small increase in acid hydrolases in the latter patients emphasizes the significance of the much larger rise observed in mucolipidosis II and III. This paper compares the level of several serum acid hydrolases in typical I-cell disease and other lysosomal storage disorders. Material Controls These comprised children hospitalized for minor illnesses, presumably not influencing their general health. On the day of sampling none of them had signs of any acute illness and none had received hormone therapy. Patients All patients with a diagnosis of a specified metabolic disorder were studied in detail clinically and chemically. In all cases except juvenile amaurotic idiocy the appropriate enzymatic diagnosis was established. Methods Sampling, preparation described earlier [ 7 ,S] .
of serum,
storage,
and enzyme
measurements
were as
Results Mucolipidosis In sera from the patients with mucolipidosis II a-mannosidase, cu-fucosidase, /3-galactosidase, N-acetylJ%glucosaminidase, a-galactosidase and fl-glucuronidase were greatly increased (Table I). The cu-glucosidase activity was normal at pH 6.0 but above normal at pH 4.5. The activity of fl-glucosidase and acid phosphatase were slightly increased. Sera from the patient with mucolipidosis III gave similar findings except that a-glucosidase was normal at pH 4.5, The total serum level of acid hydrolases was somewhat lower in the patient with mucolipidosis III than in the patients with mucolipidosis II. Mucopolysaccharidoses The activity of N-acetyl-fl-glucosaminidase and /3-glucuronidase was increased in those patients with mucopolysaccharidosis types I and II (Hurler’s and Hunter’s syndrome). @-Galactosidase showed only a slightly less marked rise. a-Mannosidase, a-fucosidase and acid phosphatase were largely normal in activity. A similar pattern, although slightly less evident, was seen in the patients with type III (Sanfilippo’s syndrome type A and B). In type IV (Morquio’s syndrome) no augmented values were noted for any of the enzymes.
ACTIVITY
155 133 220 * 180 110
96 110 130 127 101
113
100
110
16
85
84
104
125
110
106
102 -
100 -
0.21
156 *
234 ***
205 *
82
121
204 **
235 ***
1865
2502
31-250
&gaIactosidase
Statistical methods: Student’s t test was used where the individuals were three or more. Values differing significantly from the control values (* p < 0.05; ** p < 0.01; *** P < 0.001).
0.25
100%: 0.20
110
6.0 30-180
164
4.5 25-144
or-glucosidase
. 1-l. are called
160
130
130
126
Mean values for controls, expressed in /.tmol substrate split mine1 1.06 4.62 4.31 1.20
(n = 4)
102
100
103
120
728 382
300
3200
1162 681
450
6100
5.5 52-200
4.0 50-250
4.5 35-200
5.5 32-160
a-fucosidase
cu-mannosidase
IN SERUM
(range) Mucolipidosis II (n = 2) Mucolipidosis III (n = 1) Mucopolysaccharidosis I (n = 12) Mucopolysaccharidosis II (n = 4) Mucopolysaccharidoais III (n = 4) Mucopolysaccharidosis IV (n = 3) Gaucher’s disease (n = 10) Metachromatic leucodystrophy (n = 5) Juvenile amaurotic idiocy
PH Control ” = 20
ENZYME
TABLE I
0.001
-
-
178 *
82
125
98
103
390
310
60-240
p-ghlcosidase
13.5
118
170 *
318 *
120
193 **
252 ***
260 ***
i906
2101
52-200
Nacetyl-Pglucosaminidase
0.22
-
-
615
403
42-210
01galactosidase
0.08
320 ***
250 ***
278 ***
128
256 ***
352 ***
300 ***
2610
3020
5&-190
@-gIucUronidase
10.3
103
135
293 ***
82
124
108
84
180
206
61-145
Acid phosphatase
36
Gaucher’s disease The enzyme pattern in Gaucher’s disease (juvenile type) that in mucopolysaccharidosis with one important exception: was strikingly increased.
was analogous to acid phosphatase
Juvenile amauro tic idiocy Markedly increased values were found for the activity of fl-glucuronidase. Some rise was also noted for ol-fucosidase and P-galactosidase. No highly abnormal values were found for any other enzyme, including N-acetyl-@glucosaminidase.
Metachromatic
leucodystrophy
Elevated values were seen for some of the enzyme activities. The pattern was similar both to that found in juvenile amaurotic idiocy and that in the mucopolysaccharidoses types I and II.
Other lysosomai
storage disorders
The hydrolases listed in Table I have been analyzed in several other storage disorders. The following were found to exhibit normal levels of acid hydrolases in their sera: juvenile GM2-gangliosidosis, Tay-Sachs disease, juvenile Sandhoff disease (except deficiency of N-acetyl-fl-glucosaminidase in serum), Sandhoff disease (except deficiency of N-acetyl-fl-glucosaminidase), GMl-gangliosidosis (except deficiency of fl-galactosidase) and Krabbe’s disease. In each group of patients 2-3 individuals have been investigated. Discussion Inherited lysosomal storage diseases of man generally result from deficient activity of single lysosomal enzymes [ 91. The human mucolipidoses, however, represent an important group of related disorders characterized by abnormal activity of several acid hydrolases. These disorders suggest a single genetic defect, common to the final expression of several lysosomal enzymes. The primary defect in mucolipidosis II and III is not yet known, but several of the excreted lysosomal enzymes appear to lack a recognition marker necessary for normal uptake into the fibroblasts [lo]. Fibroblasts seem to ingest some of their secreted lysosomal hydrolases by a cell surface receptor that recognizes a phosphorylated carbohydrate residue, possibly phosphomannose [ 11,121. Because of the defective uptake of acid hydrolases into fibroblasts from patients with mucolipidosis II and III the enzymes in question escape into the serum in massive amounts. Recent evidence from several laboratories suggests that rapid in vivo clearance of glycoproteins (e.g. acid hydrolases) from serum is mediated by at least two distinct recognition systems. The work of Ashwell and colleagues [13] has revealed the hepatocyte-receptor-mediated plasma clearance of galactose terminal (i.e. asialo-) glycoproteins. Based on work from several laboratories [14, 151 evidence for a second pathway has emerged. The latter accommodates many lysosomal hydrolases and other glycoproteins having mannose and/or N-acetyl-glucosamine as their respective terminal sugars. This receptor would
37
appear to be connected to cells of the reticuloendothelial system [2]. The glycoproteins so cleared are degraded in the lysosomes. Thus it might be possible that the lysosomal enzymes in sera from mucolipidosis II and III are cleared less effectively than normal. This phenomenon would also contribute to the greatly increased value of serum acid hydrolases found in these patients. The increase of activity of these enzymes in the other lysosomal storage diseases would not be entirely non-specific since it was not observed in all the storage diseases studied, and the degree of elevation tended to be greater in some of the storage diseases than in others. Different enzymes were also elevated in different diseases. In mucopolysaccharidosis I, II and III a normal value was seen for acid phosphatase while in Gaucher’s disease acid phosphatase was increased. In all these diseases there was a rise in N-acetyl-P-glucosaminidase but in juvenile amaurotic idiocy and metachromatic leucodystrophy this enzyme exhibited normal levels despite raised P-galactosidase and fl-glucuronidase values. Many of the sphingolipidoses investigated (e.g. Tay-Sachs’ disease, Sandhoff disease, GMl-gangliosidosis and Krabbe disease) revealed no increase in serum acid hydrolases. A possible explanation of these findings could be that in lysosomal storage disorders, where the visceral organs are preferentially affected, resulting, for example, in hepatopathy, the diseased liver cells fail to clear acid hydrolases from the circulation. Lysosomal “overloading” might also cause a dysfunction of the lysosomal apparatus. As the asialoglycoproteins taken up by the receptors are degraded in the lysosomes a dysfunction of the latter could lead to defective clearance of asialoglycoproteins. The indications are that P-galactosidase, N-acetyl-fl-glucosaminidase and P-glucuronidase are especially sensitive to the decreased clearance as judged by the findings in the mucopolysaccharidoses and Gaucher’s disease. The increase of acid phosphatase found in sera from Gaucher’s disease might also depend upon the reticula-endothelial system and haematopoietic tissue being severely affected in this disease. Acknowledgements Skillful technical assistance was rendered by Mrs. Sonja Glans. This work was supported by the Swedish Medical Research Council (Grant No. 03X-5643) and the Medical Faculty, University of Lund. References Lightbody,
J.. Wiesmann.
Wiesman,
M.D.,
Ackerman.
P.A.
and
&kerman,
P.A.
(1968)
Den
W.R.,
Tandt,
tickerman, Chester, Thesis. 8
Eriksson,
9
Neufeld.
KBhlin,
M.A.,
Hultberg.
Lim.
10
Hickman,
11
Kaplan,
A..
Achord.
12
Ullrich.
K.,
Mersmann,
S. and
Acta
Chem.
New
Engl.
J. Lab.
Clin.
Lancet J. Med.
1, 451
285.
1090-1091
15.61-66
20.1-5
Philippart,
Masson.
N. (1971)
N. (1971)
Clin.
Paediatr.
B.,
Herschkowitz,
M. (1974)
Stand. P.K..
57,
Med.
83,403407
537-540
SjSblad,
S.
and
Kristiansson,
B.
(1977)
in
S.
Sj6blad.
Lund
Ginsburg.
E.F.,
Chim.
E. and Acta
B. and
Herschkowitz,
P. (1969)
Clin.
(1968)
PP. 91-113.
Hadorn,
F. and
Lassila,
P.A.
6..
U.,
Vassela.
B.E.,
T.W.
Neufeld, D.T. G.,
and E.F. and
Hultberg. Shapiro, (1972) Sly.
Weber.
W.S.
B. and L.J.
Ackerman.
(1975)
Biochem. (1977)
E. and Von
Ann.
P.A. Rev.
Biophys. Proc.
Natl.
Figura,
K.
(1972)
Clin.
Biochem.
Res. Acad. (1978)
Commun. Sci.
Chim.
Acta
44.357-376 USA
Biochem.
49. 74,
992-999 2026-2030
J. 170.
643-650
40,181-185
38 13
Ashwell,
G. and
14
Furbish.
F.S..
Commun. 15
Stahl,
P.D.,
1399-1403
More&
Steer,
A.G.
C.J.,
(1974)
Barranger,
Advanc. J.A..
Enzymol.
Jones,
E.A.
41.99-128 and
Brady,
R.O.
(1978)
Biochem.
Biophys.
Res.
81,1047-1053 Rodman,
J.S..
Miller,
M.J.
and
Schlesinger,
P.H.
(1978)
Proc.
Natl.
Acad.
Sci.
USA
75,
Clinica Chimica Actu, 100 (1980) 33-38 0 Elsevier/North-Holland Biomedical Press
CCA 1196
ACID HYDROLASES DISORDERS
IN SERUM FROM PATIENTS
WITH LYSOSOMAL
B. HULTBERG aq*, A. I~AKS~~N b, s. SJ~~BLAD e and P.A. ~~CKERMAN a Departments of Clinical Chemistry a and Pediatrics c, University Hospital, S-221 85 Lund (Sweden), and Department of Clinical Chemistry b, Central Hospital, S-351 85 Vtixjii (Sweden)
(Received July 3rd, 1979)
summary The activity of acid hydrolases was studied in serum from patients with mucolipidosis (II and III) and other lysosomal disorders. In mucolipidosis II and III all hydrolases examined except a-glucosidase, fl-glucosidase and acid phosphatase were greatly increased. High values for P-galactosidase were seen in mucopolysaccharidosis types I and II, Gaucher’s disease, juvenile amaurotic idiocy and metachromatic leucodystrophy. N-Acetyl-fl-glucosaminidase activity was high in mucopolysaccharidosis types I, II, III and Gaucher’s disease. The activity of /I-glucuronidase was increased in mucopolysaccharidosis types I, II and III, Gaucher’s disease, juvenile amaurotic idiocy and metachromatic leucodystrophy. Acid phosphatase had increased activity only in Gaucher’s disease. In several lysosomal storage disorders no increased values could be found. It is suggested that high values in serum from patients with lysosomal storage disorders (not including mucolipidosis II and III) may depend upon liver cell damage, which disturbs the clearing of acid hydrolases from serum.
Introduction I-cell disease (mucolipidosis II) and mucolipidosis III are especially interesting lysosomal disorders because they provide a model for the exploration of basic endo- and exocytosis phenomena of acid hydrolases. One of the main features of mucolipidosis II and III is that lysosomal enzymes are elevated in
* Correspondence Sweden.
to: Dr. BjBm Hultberg,
Dept. of Clinical Chemistry,
University
Hospital,
S-221
85 Lund.
34
the medium of cultured fibroblasts from patients with the disease and are also greatly elevated in the serum of these patients [ 1,2]. A slight increase in the activity of acid hydrolases in serum has also been found in some other lysosomal storage disorders, in patients with Gaucher’s disease [3], mucopolysaccharidosis [4,5], and in sera from patients with juvenile amaurotic idiocy [6]. The comparatively small increase in acid hydrolases in the latter patients emphasizes the significance of the much larger rise observed in mucolipidosis II and III. This paper compares the level of several serum acid hydrolases in typical I-cell disease and other lysosomal storage disorders. Material Controls These comprised children hospitalized for minor illnesses, presumably not influencing their general health. On the day of sampling none of them had signs of any acute illness and none had received hormone therapy. Patients All patients with a diagnosis of a specified metabolic disorder were studied in detail clinically and chemically. In all cases except juvenile amaurotic idiocy the appropriate enzymatic diagnosis was established. Methods Sampling, preparation described earlier [ 7 ,S] .
of serum,
storage,
and enzyme
measurements
were as
Results Mucolipidosis In sera from the patients with mucolipidosis II a-mannosidase, cu-fucosidase, /3-galactosidase, N-acetylJ%glucosaminidase, a-galactosidase and fl-glucuronidase were greatly increased (Table I). The cu-glucosidase activity was normal at pH 6.0 but above normal at pH 4.5. The activity of fl-glucosidase and acid phosphatase were slightly increased. Sera from the patient with mucolipidosis III gave similar findings except that a-glucosidase was normal at pH 4.5, The total serum level of acid hydrolases was somewhat lower in the patient with mucolipidosis III than in the patients with mucolipidosis II. Mucopolysaccharidoses The activity of N-acetyl-fl-glucosaminidase and /3-glucuronidase was increased in those patients with mucopolysaccharidosis types I and II (Hurler’s and Hunter’s syndrome). @-Galactosidase showed only a slightly less marked rise. a-Mannosidase, a-fucosidase and acid phosphatase were largely normal in activity. A similar pattern, although slightly less evident, was seen in the patients with type III (Sanfilippo’s syndrome type A and B). In type IV (Morquio’s syndrome) no augmented values were noted for any of the enzymes.
ACTIVITY
155 133 220 * 180 110
96 110 130 127 101
113
100
110
16
85
84
104
125
110
106
102 -
100 -
0.21
156 *
234 ***
205 *
82
121
204 **
235 ***
1865
2502
31-250
&gaIactosidase
Statistical methods: Student’s t test was used where the individuals were three or more. Values differing significantly from the control values (* p < 0.05; ** p < 0.01; *** P < 0.001).
0.25
100%: 0.20
110
6.0 30-180
164
4.5 25-144
or-glucosidase
. 1-l. are called
160
130
130
126
Mean values for controls, expressed in /.tmol substrate split mine1 1.06 4.62 4.31 1.20
(n = 4)
102
100
103
120
728 382
300
3200
1162 681
450
6100
5.5 52-200
4.0 50-250
4.5 35-200
5.5 32-160
a-fucosidase
cu-mannosidase
IN SERUM
(range) Mucolipidosis II (n = 2) Mucolipidosis III (n = 1) Mucopolysaccharidosis I (n = 12) Mucopolysaccharidosis II (n = 4) Mucopolysaccharidoais III (n = 4) Mucopolysaccharidosis IV (n = 3) Gaucher’s disease (n = 10) Metachromatic leucodystrophy (n = 5) Juvenile amaurotic idiocy
PH Control ” = 20
ENZYME
TABLE I
0.001
-
-
178 *
82
125
98
103
390
310
60-240
p-ghlcosidase
13.5
118
170 *
318 *
120
193 **
252 ***
260 ***
i906
2101
52-200
Nacetyl-Pglucosaminidase
0.22
-
-
615
403
42-210
01galactosidase
0.08
320 ***
250 ***
278 ***
128
256 ***
352 ***
300 ***
2610
3020
5&-190
@-gIucUronidase
10.3
103
135
293 ***
82
124
108
84
180
206
61-145
Acid phosphatase
36
Gaucher’s disease The enzyme pattern in Gaucher’s disease (juvenile type) that in mucopolysaccharidosis with one important exception: was strikingly increased.
was analogous to acid phosphatase
Juvenile amauro tic idiocy Markedly increased values were found for the activity of fl-glucuronidase. Some rise was also noted for ol-fucosidase and P-galactosidase. No highly abnormal values were found for any other enzyme, including N-acetyl-@glucosaminidase.
Metachromatic
leucodystrophy
Elevated values were seen for some of the enzyme activities. The pattern was similar both to that found in juvenile amaurotic idiocy and that in the mucopolysaccharidoses types I and II.
Other lysosomai
storage disorders
The hydrolases listed in Table I have been analyzed in several other storage disorders. The following were found to exhibit normal levels of acid hydrolases in their sera: juvenile GM2-gangliosidosis, Tay-Sachs disease, juvenile Sandhoff disease (except deficiency of N-acetyl-fl-glucosaminidase in serum), Sandhoff disease (except deficiency of N-acetyl-fl-glucosaminidase), GMl-gangliosidosis (except deficiency of fl-galactosidase) and Krabbe’s disease. In each group of patients 2-3 individuals have been investigated. Discussion Inherited lysosomal storage diseases of man generally result from deficient activity of single lysosomal enzymes [ 91. The human mucolipidoses, however, represent an important group of related disorders characterized by abnormal activity of several acid hydrolases. These disorders suggest a single genetic defect, common to the final expression of several lysosomal enzymes. The primary defect in mucolipidosis II and III is not yet known, but several of the excreted lysosomal enzymes appear to lack a recognition marker necessary for normal uptake into the fibroblasts [lo]. Fibroblasts seem to ingest some of their secreted lysosomal hydrolases by a cell surface receptor that recognizes a phosphorylated carbohydrate residue, possibly phosphomannose [ 11,121. Because of the defective uptake of acid hydrolases into fibroblasts from patients with mucolipidosis II and III the enzymes in question escape into the serum in massive amounts. Recent evidence from several laboratories suggests that rapid in vivo clearance of glycoproteins (e.g. acid hydrolases) from serum is mediated by at least two distinct recognition systems. The work of Ashwell and colleagues [13] has revealed the hepatocyte-receptor-mediated plasma clearance of galactose terminal (i.e. asialo-) glycoproteins. Based on work from several laboratories [14, 151 evidence for a second pathway has emerged. The latter accommodates many lysosomal hydrolases and other glycoproteins having mannose and/or N-acetyl-glucosamine as their respective terminal sugars. This receptor would
37
appear to be connected to cells of the reticuloendothelial system [2]. The glycoproteins so cleared are degraded in the lysosomes. Thus it might be possible that the lysosomal enzymes in sera from mucolipidosis II and III are cleared less effectively than normal. This phenomenon would also contribute to the greatly increased value of serum acid hydrolases found in these patients. The increase of activity of these enzymes in the other lysosomal storage diseases would not be entirely non-specific since it was not observed in all the storage diseases studied, and the degree of elevation tended to be greater in some of the storage diseases than in others. Different enzymes were also elevated in different diseases. In mucopolysaccharidosis I, II and III a normal value was seen for acid phosphatase while in Gaucher’s disease acid phosphatase was increased. In all these diseases there was a rise in N-acetyl-P-glucosaminidase but in juvenile amaurotic idiocy and metachromatic leucodystrophy this enzyme exhibited normal levels despite raised P-galactosidase and fl-glucuronidase values. Many of the sphingolipidoses investigated (e.g. Tay-Sachs’ disease, Sandhoff disease, GMl-gangliosidosis and Krabbe disease) revealed no increase in serum acid hydrolases. A possible explanation of these findings could be that in lysosomal storage disorders, where the visceral organs are preferentially affected, resulting, for example, in hepatopathy, the diseased liver cells fail to clear acid hydrolases from the circulation. Lysosomal “overloading” might also cause a dysfunction of the lysosomal apparatus. As the asialoglycoproteins taken up by the receptors are degraded in the lysosomes a dysfunction of the latter could lead to defective clearance of asialoglycoproteins. The indications are that P-galactosidase, N-acetyl-fl-glucosaminidase and P-glucuronidase are especially sensitive to the decreased clearance as judged by the findings in the mucopolysaccharidoses and Gaucher’s disease. The increase of acid phosphatase found in sera from Gaucher’s disease might also depend upon the reticula-endothelial system and haematopoietic tissue being severely affected in this disease. Acknowledgements Skillful technical assistance was rendered by Mrs. Sonja Glans. This work was supported by the Swedish Medical Research Council (Grant No. 03X-5643) and the Medical Faculty, University of Lund. References Lightbody,
J.. Wiesmann.
Wiesman,
M.D.,
Ackerman.
P.A.
and
&kerman,
P.A.
(1968)
Den
W.R.,
Tandt,
tickerman, Chester, Thesis. 8
Eriksson,
9
Neufeld.
KBhlin,
M.A.,
Hultberg.
Lim.
10
Hickman,
11
Kaplan,
A..
Achord.
12
Ullrich.
K.,
Mersmann,
S. and
Acta
Chem.
New
Engl.
J. Lab.
Clin.
Lancet J. Med.
1, 451
285.
1090-1091
15.61-66
20.1-5
Philippart,
Masson.
N. (1971)
N. (1971)
Clin.
Paediatr.
B.,
Herschkowitz,
M. (1974)
Stand. P.K..
57,
Med.
83,403407
537-540
SjSblad,
S.
and
Kristiansson,
B.
(1977)
in
S.
Sj6blad.
Lund
Ginsburg.
E.F.,
Chim.
E. and Acta
B. and
Herschkowitz,
P. (1969)
Clin.
(1968)
PP. 91-113.
Hadorn,
F. and
Lassila,
P.A.
6..
U.,
Vassela.
B.E.,
T.W.
Neufeld, D.T. G.,
and E.F. and
Hultberg. Shapiro, (1972) Sly.
Weber.
W.S.
B. and L.J.
Ackerman.
(1975)
Biochem. (1977)
E. and Von
Ann.
P.A. Rev.
Biophys. Proc.
Natl.
Figura,
K.
(1972)
Clin.
Biochem.
Res. Acad. (1978)
Commun. Sci.
Chim.
Acta
44.357-376 USA
Biochem.
49. 74,
992-999 2026-2030
J. 170.
643-650
40,181-185
38 13
Ashwell,
G. and
14
Furbish.
F.S..
Commun. 15
Stahl,
P.D.,
1399-1403
More&
Steer,
A.G.
C.J.,
(1974)
Barranger,
Advanc. J.A..
Enzymol.
Jones,
E.A.
41.99-128 and
Brady,
R.O.
(1978)
Biochem.
Biophys.
Res.
81,1047-1053 Rodman,
J.S..
Miller,
M.J.
and
Schlesinger,
P.H.
(1978)
Proc.
Natl.
Acad.
Sci.
USA
75,