Amazing A-beta All Stars

…what serious money and great talent tell us about b -amyloid

P. Thiyagarajan, et al.— Argonne National Laboratory & University of Chicago Applied Crystallography Article J. Appl. Cryst., 33, 535-539 (2000) This article describes the fibril structure of a sub-peptide of A b , sometimes with the attach-ment of a blocking polyethylene glycol.

The idea is to figure out whether and how an attachment to the A b will affect the fibril formation. H 2 N A b 10-35 PEG

SANS = Small Angle Neutron Scattering Argonne National Laboratory (near Chicago) Radially-averaged intensity profile Detector with time of flight capabilities, can measure over a wide range of scattering vectors simultaneously Sample 0.5 Å to 15 Å neutrons in each pulse Pulsed neutron source

SANS

I

(

q

) Radially-averaged intensity profile Destructive interference between neutrons scattered from different parts of the Amyloid fibrils causes the intensity to decline with angle. The specific pattern of that decline can be interpreted in terms of fibril thickness and mass per unit length.

q

Interferences due to fibrillar shape can be “taken out” of the data by multiplying intensity by scattering vector magnitude, q . The remaining decline in intensity is due to finite fibril thickness.

Rod-Modified Guinier Plot—Slope Gives Fibril Cross-sectional Radius

R

g,c

Findings

Fibril thickness depends strongly on pH.

Fibril-fibril associations are less at low pH (even though, from our experience, such conditions lead to rapid self-assembly of A b ).

Fibrils are thicker when PEG is present (by 23 – 30 Å) PEG is located along the periphery of the fibril.

Interdisciplinary Considerations

Think what goes into such an experiment • Someone to build/program SANS (several $M) • Someone to make peptide • Someone to attach PEG • Someone to collect & analyze data • Someone with the intellectual lead to direct project, set standards, monitor competitors & write the results. • Nine (9) authors in all!

Robert Tycko National Institutes of Health, Maryland Current Topics Article (i.e., review) Biochemistry, 42(11), 3151-3159 (2003) If I wrote one article like this one in my whole life, that would represent a great career. General purpose of the article: review structural methods, especially solid state NMR, for determining b -amyloid structure.

Amyloid does not crystallize, so crystallography won’t suffice.

The information required exceeds what small angle X-ray and Small angle neutron scattering can provide. No choice but to haul out a big NMR (and serious labeling, simulation, etc.).

13 C labeling off-center will produce different dipole-dipole coupling for Free peptides Tycko Figure 2 Various derivatives (circles) and simulated for different distances (lines) “These data indicate an in register, parallel alignment….”

13 C 13 C exchange using a 9.4T NMR indicates which specifically labeled residues are closely coupled.

Multiple-quantum NMR absorptions require proximal nuclei. “An

n

-quantum 13C NMR signal is observable only if at least

n

13C nuclei are sufficiently close in space to be linked by a network of couplings with coupling constants

d ~

1/ t MQ ” Again, simulations are performed to show that parallel alignment of the peptides occurs. Figure 3

Figure 6 Proposed Model of Amyloid Each molecule has 2 beta strands, red and blue Hydrophobic Polar Positively charged

EM provides less detail, but it is so much easier!

Helmuth M Ö hwald & Coworkers Max Planck Institute for Colloids and Interfaces Golm, Germany This group is devoted to surface studies.

One mechanism of nano-organization and self assembly that we have not discussed is organization of nanostructures at surfaces.

So…here is a good example concerning how A b organizes near the air-water interface. Air is hydrophobic (despite the humidity in Baton Rouge or Bangkok!) so this is a model for interaction with other hydrophobic surfaces in the body—cell walls, some protein surfaces, etc. That kind of interaction is a little more complex—a colleague studies it back at LSU.

Reflection Infrared Absorption

Langmuir Trough

0.0015 Wilhelmy plate surface tension measurement Movable barrier

Surface Reflection Absorption Infrared “ The central message of this work is that a stable b sheet-enriched state of the amyloid is formed at the air water interface, in contrast to the initial bulk solution containing high a helix/random coil….”

Stability

It was also found that the reflection IR spectra were stable to several compression/dilation cycles.

In the plane of the air-water interface there exist antiparallel b -sheet structures. Certain alignments of the self-assembled structure could be ruled out, but the essential information is that the surface induces b -sheet formation. For structural details, the Tycko work is better.

Alignment

Certain alignments of the self-assembled structure could be ruled out, but the essential information is that the surface induces better.

b -sheet formation. For structural details, the Tycko work is

Kim & Lee Fluorescence Nanoscopy Laboratory Department of Chemistry Ewha Womens University Seoul, Korea Nanomedicine: can a Buckeyball cure Alzheimer’s?

My colleagues laughed at this paper.

Afterall, we are seriously about making peptide inhibitors at LSU, and everyone knows (or thinks they know) that the answer will be a peptide.

But let’s have a look.

Buckey inhibitor?

Fluorescence data shows that the selected Fullerene suppresses A b aggregation. Compare to Melatonin, a circadean hormone.

IC 50 values (inhibitory concentration for 50% reduction) Fullerene works about as good as anything else!

Point of Discussion

Scattering and NMR provide indirect evidence for the structure. That is the most detailed info we have, but….

In the future, what will be the role of NMR, scattering and other techniques? If we are to build Erick Drexler’s nano-robots, won’t they have nano-vision? Tim Lodge story.

What’s Next?

Advancements in Cryo-TEM may give soft matter microscopy almost the power that solid state enjoys (i.e., sub-nanometer) Jimenez et al.

EMBO Journal 18(4), 815-821 (1999) “890 cut-out repeats* were iteratively aligned by a multivariate statistical analysis.” Then the images are manipulated in 3D *Image sections Microscopy at 25 Å resolution

Conclusion

• • • • Disease-inspired inventions and innovations to study nanometer-scale objects will continue to be important as non-biological nanoscience develops. With apologies to Richard Feynman, there is always room at the interfaces !

Some interesting interfaces biological-materials fundamental-applied theory-imaging nano-micro