RESOLVING THE HIGH-RESOLUTION ARCHITECTURE, ASSEMBLY AND FUNCTIONAL REPERTOIRE OF BACTERIAL SYSTEMS BY IN VITRO ATOMIC FORCE MICROSCOPY - Life at the Nanoscale: Atomic Force Microscopy of Live Cells - page 85

play important roles in microbial attachment, dispersal and pathogenesis. 15
We have proposed

spore
coat rodlets and the amyloid rodlets found on other bacterial and fungal
spores suggests that

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that the structural similarity of B. atrophaeus

rodlets have an amyloid structure. AFM
characterization of the nanoscale properties of individual amyloid ibrils
has revealed that these self-assembled structures can have a strength and
stiffness comparable with structural steel. 53 The extreme physical, chemical
and thermal resistance of

Bacillus

spores suggests that evolutionary forces
have captured the mechanical rigidity and resistance of these amyloid self-
assembling biomaterials to structure the protective outer spore surface.
Structural studies of amyloids have identiied an array of possible rodlet
assemblies, each consisting of several (2 or 4) individual cross-β sheet ibrils,
which are often helically intertwined. 15 The number of ibrils determines
the diameter of the rodlet. Most amyloids resulting from protein folding
diseases, and some naturally occurring amyloids, form individual ibrils or
disorganized rodlets networks.
In spore coats of

Bacillus

, the higher-order rodlet structure is
organized as one major domain of parallel rodlets covering the entire spore
surface. 3 Rodlet domain formation requires the periodic bonds in the rodlet
direction (“parallel bonds”) as well in the direction perpendicular to it
(“perpendicular bonds”). 54 In the case of amyloid-like rodlets, the

B. atrophaeus

-rodlet,
parallel bonds are known and consist primarily of hydrogen bonds associated
with the cross-β sheets that form the backbone of the rodlet ibrils. However,
the nature of the perpendicular bonds,

intra

i.e. the inter -rodlet bonds that keep

the rodlets tightly packed, is unknown.
On the basis of these rodlet features, one might expect that during
germination individual rodlets would detach or erode, leaving a striated
pattern

to the rodlet direction were observed ( Fig. 4.9 ) , and 2-3 nm wide ibrils
perpendicular

parallel to the rodlet direction. Surprisingly, striations perpendicular

to the rodlet direction ( Fig. 4.9e ) were the culmination
product of coat degradation. This result indicates that during germination,
perpendicular rodlet bonds are stronger, or are more resistant to hydrolysis,
than bonds parallel to the rodlet direction. Second, and most surprisingly,
these perpendicular structures facilitate the formation of 200-300 nm long
ibres

to the rodlet direction.
It is unclear how microbial amyloid ibres form these perpendicular
structures. One possibility is that during the formation of the rodlet layer,
both intra-rodlet parallel bonds and inter-rodlet perpendicular bonds form,
similar in strength and leading to tightly packed rodlets domains held together
by a checkerboard-like bonding pattern. During germination, the intra-rodlet
parallel bonds are hydrolyzed, while the inter-rodlet perpendicular bonds
remain intact over longer time periods. Spore coat hydrolytic enzymes
could target a speciic residue or structure (in this case, that of the cross-β

perpendicular