Fungal Polysaccharides

285 Pages·1980·4.13 MB·English

by  Paul A. Sandford and Kazuo Matsuda (Eds.)

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Fungal Polysaccharides
Paul A. Sandford, EDITOR
Kelco, Division of
Merck and Company, Incorporated
Kazuo Matsuda, EDITOR
Tohoku University
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19 Based on a symposium
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Honolulu, Hawaii, April 4-5, 1979.
126
ACS SYMPOSIUM SERIES
AMERICAN CHEMICAL SOCIETY
WASHINGTON, D. C. 1980
1 Library of Congress CIP Data
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w Fungal polysaccharides.
6.f (ACS symposium series; 126 ISSN 0097-6156)
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0 Includes bibliographies and index.
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98 1. Microbial polysaccharides—Congresses. 2. Fungi
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k- —Composition—Congresses. 3. Cell membranes—
1/b Composition—Congresses.
02 I. Sandford, Paul Α., 1939- . II. Matsuda,
0.1 Kazuo, 1922- . III. American Chemical Society.
oi: 1 DChiveimsiiocna l oSfo cCieatryb. ohDyidvirsaitoen Cohfe mAgisrtircyu. ltuIVra.l aAnmd eFriocoadn
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0 | Chemistry. V. ACS/CSJ Chemical Congress, Hono­
8 lulu, 1979. VI. Series: American Chemical Society.
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0, 1 ACS symposium series; 126.
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e QR92.P6F86 589.2'0419282 80-10639
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Robert A. Hofstader Gunter Zweig
FOREWORD
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6.f The ACS SYMPOSIUM SERIES was founded in 1974 to provide
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k-1 EST CHEMISTRY SERIES except that in order to save time the
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doi: of the Series Advisory Board and are selected to maintain the
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PREFACE
Most fungi produce some type of polysaccharide that can be found in
cell membranes or walls and as exocellular components. The role
of these polysaccharides is not well understood, but in some cases they
function as structural components of the fungal cell wall. This does not
mean that fungal polysaccharides do not have important roles. For ex
ample, fungal polysaccharides are involved directly in host-pathogen
interactions between fungi in mammals and plants. Also, surface exo
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00 cellular polysaccharides of industrially useful fungi (yeast in particular)
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6. have been studied in great detail in order to determine their role in vari
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01 ous specific applications.
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8 Like bacterial polysaccharides, fungal polysaccharides of several
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k- types have been discovered that vary widely in their sugar composition,
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1/ linkage types, molecular parameters, and physical properties.
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1 This volume focuses on various aspects of fungal polysaccharide
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oi: 1 research that are currently being carried on throughout the world.
d Therefore, topics in this volume vary from the genetics of polysaccharide
80 | biosynthesis to host-pathogen interactions and from immunochemistry to
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0, 1 the chemical and physical characterization of specific and interesting
e 1 fungal polysaccharides that have industrial utility.
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ate: Kelco, Division of PAUL A. SANDFORD
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Sendai, 980 Japan
October 23, 1979
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1
Genetics of Yeast Mannoprotein Biosynthesis
CLINTON E. BALLOU
Department of Biochemistry, University of California, Berkeley, CA 94720
In this paper I intend to summarize what we know about yeast
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0 mannoprotein structure, what features of the biosynthetic pathway
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26. gaining this information, and what is known about the genetic
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k-1 Yeast Mannoprotein Structure
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02 Prior to 1965, the limited information available was due
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0. mainly to the methylation studies of Haworth et al. (1) which in­
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oi: dicated that yeast mannan was a highly-branched polysaccharide
d with 1->2, 1->3 and 1->6 linkages, in no specific order. Dramatic
0 | developments have occurred during the last 10 years that have
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19 revolutionized our understanding of the structure of this very
10, complex glycoprotein we now call a mannoprotein. The first im­
ne portant development was the acetolysis procedure, first applied
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ate: in our laboratory (3-10), that allowed selective cleavage of the
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n 1->6 linkages in the mannan component of the glycoprotein and fa­
atio cilitated the isolation of oligosaccharide fragments whose struc­
c tures could then be established by classical methods. From a
ubli comparison of the acetolysis oliogsaccharide patterns (Figure 1)
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from different mannoproteins (6), it was clear that the mannan
structures were species-specific (11).
The second important step in elucidation of mannoprotein
structure was the discovery of an enzyme with the ability to re­
move the mannan sidechains selectively so that the backbone
structure could be studied (12). As a consequence, several yeast
mannans, although not all, have been shown to have a linear α1->6-
linked backbone (13, 14, 15). As a corollary, it follows that
the acetolysis oligosaccharides represent sidechain fragments of
the mannan, and by their characterization we were able to arrive
at schematic structures for the mannan chains (Figure 2).
These two developments facilitated a third; namely, the elu­
cidation of the immunochemistry of the yeast mannoproteins.
0-8412-0555-8/80/47-126-001$05.00/0
© 1980 American Chemical Society
2 FUNGAL POLYSACCHARIDES
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80- Figure 1. Acetolysis oligosaccharide patterns from different yeast mannopro­
19 teins (11)- The products from the reaction, which was carried out according to Kef. 6,
k- were separated on a Bio-Gel P-2 column (2 X 200 cm) by elution with O.J M acetic acid.
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021/ M tot eMtra5 oasree mphaonsnpohsaet et,o amnda nMnfoipNeAnct aoiss eN, -Mac3Pet yislg mluacnonsoatmriionsee l ipnhkoesdp thoa mtea, nMn^oPte itsr amosaen. no-
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e 1 ^2 j>2 φ2 ^2 ^2 φ2
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ation αΜ+ 3 αΜ+ 3
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Saccharomyces cerevisiae Kluyveromyces lactis
αΜ^6αΜ^6αΜ^6αΜ^6αΜ^ αΜ^Μ+6αΜ+6αΜ+6αΜ+
^2 ^2 ^2 ^2 ^2 ^2 j.2 j.2
Ρ+6αΜ αΜ αΜ αΜ αΜ αΜ αΜ αΜ
+ f2 +2 f2
αΜ αΜ αΜ αΜ aGNAc+2aM αΜ αΜ
αΜ αΜ αΜ αΜ αΜ
Figure 2. Structures for the mannoproteins whose acetolysis patterns are shown
in Figure 1. The structures illustrate the nature of the backbone and sidechains
but they are not intended to indicate the order or number of sidechains.
1. BALLOU Yeast Mannoprotein Biosynthesis 3
Rabbit antisera raised against intravenously injected whole yeast
cells give strong precipitin reactions with isolated mannopro­
teins (l6) and the homologous reactions can usually be inhibited
by acetolysis fragments from the mannan (ΓΓ, l8_, 19., 20) . Such
studies revealed that the immunodominant structures of the yeast
cell surface are the mannoprotein sidechains, and that some side-
chains are more immunogenic than others (Figure 3).
The discovery that the mannan component of the yeast cell
was the major surface determinant produced important dividends,
for it suggested a procedure by which mutants with altered manno­
proteins might be obtained. The procedure involved the selection,
from a mutagenized yeast culture, of variants that no longer were
agglutinated by antiserum prepared against the parent cells (21).
Because the only phenotypic expression of such mutants is a
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ch acterization of the mutants required extensive structural analy­
26. sis of the cell walls (22_). The screening for cell surface
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0-0 mutants, as contrasted with selection or enrichment, can be done
8 with antisera of different specificities (23) or with the Alcian
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k-1 Blue dye binding assay {2k) that will detect the presence or ab­
1/b sence of phosphate in the mannan. A recent improved procedure,
02 reported at this meeting (25), describes the use of a fluorescent
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0. lectin for the selection and enrichment of yeast mannoprotein
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oi: mutants by a fluorescence-activated cell sorter. This procedure
d should be of general use if fluorescent antibodies can be em­
80 | ployed in place of lectin.
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0, 1 The various mannoprotein mutants will be discussed in a
1 later section of this report, but one mutant must be considered
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n now because it has played an extremely important role in eluci­
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at mutant, which is characterized by an acetolysis pattern that re­
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n veals mostly mannose and indicates that the mannoprotein has a
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ati predominantly unbranched al->6-linked backbone structure (2l).
blic Studies by Dr. Tasuku Nakajima, while a postdoctoral visitor in
Pu my laboratory, demonstrated that the small amount of branching
still detected in the mannoprotein represented the presence of a
core structure that linked the main mannan chain to the protein
(Figure k) {26). A very important conclusion of this study is
that yeast mannoproteins and mammalian and plant glycoproteins
have identical asparagine-linked core carbohydrate structures
(27), and that this core structure is modified in yeast by the
addition of a polymannose outer chain. A final point of interest
is the fact that the yeast core structure, even in a homogeneous
glycoprotein, is heterogeneous (28).
Yeast Mannoprotein Biosynthesis
A number of early studies on mannan biosynthesis (29_, 30)
were done without a clear idea of the structure of the substance
and, consequently, were difficult to interpret. With the