Interactions of algal spores and diatoms with mixed synthetic peptide SAMs.
Interactions of algal spores and diatoms with mixed synthetic peptide SAMs. T. Ederth*, P. Nygren, T. Ekblad, M. Östblom, B. Liedberg Division of Molecular Physics, Department of Physics, chemistry and biology, Linköpings Universitet, SE-581 83 Linköping, Sweden, * [email protected] M. E. Pettitt, M. E. Callow, J. A. Callow The University of Birmingham, School of Biosciences, Birmingham B15 2TT, UK Advanced nanostructured surfaces for the control of biofouling http://www.ambio.bham.ac.uk/ The problem – marine biofouling Marine biofouling incurs additional costs and environmental impact on the operation of vessels, heat exchangers, fishing equipment, desalination plants and many other facilities, by reduced efficency and increased fuel consumption. There are about 2500 fouling marine organisms, using a broad range of settlement strategies and bioadhesives. We study the interactions between surfaces and marine organisms in terms of physicochemical cues, in order to understand settlement behaviour and ultimately to improve anti-fouling properties of surfaces. Mixed peptide SAMs A synthetic cationic arginine-rich peptide is used to form mixed SAMs on gold substrates (via a thiol linker), with a shorter spacer molecule. IRAS, wetting and ellipsometry data demonstrate that the peptides form organised monolayers. Mol-% ArgTyr Thickness (Å) Contact angles (θ θa) 0% 13.1 41° 5% 16.5 46° H2N ArgTyr O 20% 16.8 47° 50% 21.5 51° 75% 23.3 52° 100% 26.2 44° O H N H2 N O HS NH O H N N H O N H H2 N O H N N H H2N HS H2N O O H N N H O H N O N H HO O H N NH HN O HO O Gly3 spacer NH HN O HO Thiol ”anchor” NH HN ( θr < 10 for all mixtures ) Biological tests H2N HN N H O H N O N H 1 Normalized peak area Absorbance We have studied the attachment of a major ship fouling alga (Ulva) and a diatom (Navicula) to the mixed peptide SAMs. Solution Mol-% ArgTyr 100% 75% 50% 20% 5% 0.8 0.6 0.4 0.2 0% 1800 Spores Ulva linza Navicula perminuta Ulva plants release spores which swim with the aid of flagella. As a suitable surface for settlement is located, a physiological change into a plant is started. Ulva spores lack cell walls, and expose the plasma membrane to the environment, while the Navicula cells are encased within a silica cell wall. The peptide SAMs were incubated in Ulva spores or Navicula cells for 1 h, whereupon the density of settled (attached) cells was determined. Incubation assay -2 Spores (mm ) 2000 ArgTyr Arg Tyr Gly 1500 1000 500 0 0 20 40 60 80 100 Peptide concentration (µM) Up to 20 µM, free ArgTyr peptides in solution stimulate the settlement of Ulva spores. Above 20µM, spores lysed (died). No such effects were observed for the free amino acids of which the ArgTyr peptide is composed. 0 1000 0 20 40 60 80 100 % ArgTyr in solution FTIR data for mixed ArgTyr/spacer SAMs. Peak areas within the shaded region were used to estimate the surface fraction of the ArgTyr peptide for different solution fractions (right). Settlement results The density of settled Ulva spores increases with bound ArgTyr content but many of the 'settled' spores retained their flagella indicating that they had not gone through the normal settlement process; these spores probably died. Navicula cells, however, are unaffected by the presence of the ArgTyr peptide; cell attachment is not altered and no cells died. This difference is probably due to the diatom cell wall preventing the bound ArgTyr peptide from contacting the plasma membrane. 300 Spores per mm2 Plant 1600 1400 1200 Wavenumber (cm-1) Total Ulva spores 250 5 µm With flagella 200 Dead spores 150 100 50 0 0 5 20 50 75 Mol-% ArgTyr on surface 100 Normally settled (A) and ‘stuck’ (B) spores, with flagella still attached. Summary: The ArgTyr peptide interacts specifically with Ulva cell membranes, promoting adhesion to a surface, but also preventing it from going through its normal settlement physiology, and ultimately causing cell death.