13 oct. 2005gw canters leiden univ1. scanning probe microscopy (spm) atomic/scanning force...

13 Oct. 2005 GW Canters Leiden Univ 1

I I I . Imaging

A. SPM

B. Fluorescent labeling

Scanning Probe Microscopy (SPM)

Atomic/scanning Force Microscopy

Scanning Tunnelling Microscopy

AFM resolution

Chris Chan, MetProt Group, private comm. 2004

Mica 5nm Au nanoparticle

Chris Chan, MetProt Group, private comm. 2004

2 & 5nm Au particles on mica, TM in water

Chris Chan, MetProt Group, private comm. 2004

5nm Au particles on HOPG

Chris Chan, MetProt Group, private comm. 2004

13 Oct. 2005 GW Canters Leiden Univ 7

J.Davis et al., Chem Communic. (2003) Advance pub.

13 Oct. 2005 GW Canters Leiden Univ 8

J.Davis Pers. Communic. (2003)

13 Oct. 2005 GW Canters Leiden Univ 9

Forcing current through a single metalloprotein

Fluid conductance

J.Davis Pers. Communic. (2003)

13 Oct. 2005 GW Canters Leiden Univ 10

J.D. Davis, D. Bruce, G.W. Canters, J. Crozier, H.A.O. Hill (2003) Chem. Commun. 576-577.

Height distribution was found to be the same for the wt and K27C azurins, while the height of S118C is 5 - 8 Å lower

AFM permits to distinguish between the “standing” and the “lying” azurin molecules.

Electrochemical characteristics of the azurin monolayers.

Half wave potential is defined as (Epa–Epc)/2 vs. SCE

Azurin WT Azurin K27C Azurin S118C

E1/2*(mV vs. SCE) 165–175 165–175 120–145

ket/s–1 300–400 210–320 300–570

BUT: the electron transfer rate is independent of the distance between the Cu and the electrode surface

Is it possibly an effect of heterogeneous binding?

13 Oct. 2005 GW Canters Leiden Univ 11

Flavodoxin

13 Oct. 2005 GW Canters Leiden Univ 12

Flavodoxin N106C dimer on mica~0.1ug/ml in 20mM MES pH6TM in air

STM images of tip-induced dissociation of K27C azurin dimer, (a) before and (b) after application of a voltage pulse. Scanning conditions were bias voltage 0.1 V, current 0.1 nA, temperature 20±1 °C, relative humidity 35±5%. Halliwell et.al. 2004

Chris Chan, MetProt Group, private comm. 2004

The trimeric structure of copper-containing NiR

Monomer Trimer

Nir on template stripped gold10ug/ml in 20mM MES, pH 6TM in air

Chris Chan, MetProt Group, private comm. 2004

13 Oct. 2005 GW Canters Leiden Univ 15

Nir on template stripped gold

100ug/ml in 20mM MES, pH 6

TM in air

Chris Chan, MetProt Group, private comm. 2004

1ug/ml Nir M94C, polylysine on mica

TM in 20mM MES, pH6Chris Chan, MetProt Group, private comm. 2004

13 Oct. 2005 GW Canters Leiden Univ 17

I I I . Imaging

A. SPM

B. Fluorescent labeling

13 Oct. 2005 GW Canters Leiden Univ 18Nature Reviews Mol. Cell Biol. 3(2002)906

JACS 124(2002)6063

PNAS 101(2004)523

13 Oct. 2005 GW Canters Leiden Univ 19

Nature Reviews Mol. Cell Biol. 3(2002)906