Theoretical and experimental studies of  self-assembling protein and lipid systems

Tamás Beke-Somfai

Research Centre for Natural Sciences, Hungarian Academy of Sciences

1117

Budapest, Magyar tudósok krt.2

Summary

In a wide range of biological activities, from cell locomotion to membrane transport, Nature

deploys numerous sophisticated molecular machines which have become highly

optimized for performance and controllability. These assemblies are often

composed of multiple separate components which gather for various specific

purposes. Rational design and engineering of similarly complex biosystems is a

very exciting field with a potential to dramatically alter future’s medicine or

industrial biochemistry [1, 2]. However, to overcome major challenges in areas

such as design of artificial enzymes, or membrane active designed compounds

mimicking natural ones, the precise understanding of their mechanisms especially

of their key steps is required. Here I will focus on two examples where it is

challenging to gain insight to such mechanistic details.

1. FoF1 ATP synthase is interesting as a model

system: a delicate molecular machine synthesizing or hydrolyzing ATP utilizing

a rotary motor. Isolated F1 performs hydrolysis with a rate very

sensitive to ATP concentration. Experimental and theoretical results show that

at low ATP concentrations ATP is slowly hydrolyzed in the so called tight

binding site, whereas at higher concentrations the binding of further ATP

molecules induce rotation of the central g-subunit thereby

forcing the site to transform via subtle conformational changes into a loose

binding site, in which hydrolysis occurs faster. By a combination of

theoretical approaches we addressed how large macromolecular rearrangements may

manipulate 1?-scale rearrangements in the active site and how the reaction rate

changes as a consequence [3]. Simulations reveal that in response to g-subunit position,

the active site conformation is fine-tuned mainly by small a-subunit changes

[4]. It is hoped that in the future the design of bioinspired complex systems

arrives to the age where fine-tuning and precise control on desired processes

can be achieved

2. In the recent decades, development of resistance by bacteria to

antibiotics makes better understanding of antimicrobial mechanisms increasingly

important. Toxic oligomers of antimicrobial peptides (AMPs) may assemble into

hydrophilic or lipophilic complexes and exert their toxicity in a higher level

aggregate form. However, this mechanism is not understood, greatly hindering

rational development of similar compounds.

In this part of the presentation an overview is given on our recent

studies related to both natural and non-natural peptide oligomer assemblies and

their aggregates when associated with organic small molecules. We have

experienced several interactions resulting in induced conformational changes

for these compounds. Several of the observed secondary structures are rather

different from those regularly obtained for well studied AMPs indicating that

the action mechanism of these compounds may be different when exerting their

toxicity in in vivo conditions in presence of a complex multicomponent

environment. Also, I aim to describe new methods available in our laboratory

which are capable to address membrane systems in solution phase. In particular

I will focus on polarized light spectroscopy and on Linear Dichroism coupled to

a Couette Flow-cell (Flow-LD). By today, flow-LD can be used to characterize bicellar

systems or induce lipid bilayer fusion to test mechanisms related to cell

fusion or lipid bilayer mixing [5-7]

[1] Senes Curr. Op. Struct. Biol. 2011, 21, 460-466

[2] Jiang et al. Science, 2008, 319, 1387-1391

[3] Beke-Somfai, et al. Proc. Natl. Acad. Sci. USA, 2011, 108, 4828-4833

[4] Beke-Somfai, et al. Proc. Natl. Acad. Sci. USA, 2013, 6, 2117-2122

[5] Nordén et al. (2010), Linear Dichroism and Circular Dichroism. A Textbook on Polarized-Light Spectroscopy.

[6] Kogan et al. Langmuir, 2014, 30, 4875-4878

[7] Rocha et al. Langmuir, 2016, 32, 2841-2846