Biomedical Engineering Reference
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that the aggregation of CS proteins was an essential part of natural cellulose I biogenesis.
The aggregation of multiple polymerization sites in the plasma membrane allows the
synthesis of numerous ß1,4-glucan chains in a spatially compact area, which facilitates
cellulose I crystallization with parallel chain conformation by reducing the chances of
individual chain folding. FF-TEM imaging of unusual CSC arrays during secondary
wall deposition in a green alga,
Micrasterias denticulata
, showed that multiple CSCs
could contribute to the biogenesis of a single cellulose nanofibril, the dimensions of
which were determined by the geometric arrangement of the CSCs. The density of
CSCs is also higher in xylem cells synthesizing cellulose-rich secondary walls, which
have thicker cellulose nanofibrils compared to cells synthesizing primary walls.
Although these cell biological observations made a compelling case that CSCs did
facilitate cellulose nanofibril biogenesis, it was not until 1990 and afterwards that
this hypothesis was proven through identification of
CS
genes. After identification in
1990 of a prokaryote
CS
gene in
Gluconacetobacter xylinus
(then called
Acetobacter
xylinum
), plant
CesA
genes encoding structurally similar proteins were identified in 1996
among genes expressed during secondary wall deposition in cotton fiber. An essential
role for CesA proteins in plant cellulose synthesis was demonstrated genetically in
1998 through mutant analysis in Arabidopsis. Although there was substantial sequence
divergence between major lineages, all of the CS proteins were classified as UDP-
glucose: 1,4-ß-D-glucosyltransferase enzymes in glycosyltransferase family 2 (GT2).
Glycosyltransferases are enzymes that transfer sugar moities from activated sugar
donors, thereby forming a glycosidic bond in a short oligosaccharide or long chain
polysaccharide. The CSs are large membrane spanning proteins, for example, the
Arabidopsis CesA proteins range from 985 to 1088 amino acids and
∼
110-120 kDa
mass. The fundamental advances summarized above set the stage for probing more
deeply into the nanoscale structure and function of the CSC during the last decade.
2.3
CesA Protein is a Major Component of the Plant CSC
Land plants as well as their charophyte algal progenitors (collectively called the strepto-
phytes) have a rosette-type CSC with hexagonal symmetry. In 1999, immunolabeling in
conjunction with FF-TEM showed that the rosette CSC contained CesA protein (6). The
rosette region of the CSC is about 25 nm in diameter and contains six intramembrane
particles. Each of the six subunits is
8nmindiameter,reflectinganaggregateofCesA
proteins (Figure 2.1). It is important to recognize that the rosettes visualized by FF-TEM
are only the signatures on the cleaved inner leaflet of the plasma membrane of aggregates
of CesA transmembrane helices (TMH) traversing the membrane. The catalytic loop is
believed to exist in the cytoplasm based on protein structure prediction algorithms and
the cytoplasmic location of substrate UDP-glucose (7). The predicted proximity of adja-
cent TMH suggests that very little of the protein exists on the exoplasmic surface of the
plasma membrane.
Figure 2.2 shows schematic diagrams of the domain structure of CS proteins of organ-
isms from diverse lineages (
Gluconacetobacter xylinus
(a eubacterium),
Anabaena vari-
abilis
(a photosynthetic cyanobacterium),
Dictyostelium discoideum
(a protistan social
amoeba), and
Gossypium hirsutum
(a vascular plant)).
∼
All known CS proteins share
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