Precisely tuneable energy transfer system using peptoid helix-based molecular scaffold

The energy flow during natural photosynthesis is controlled by maintaining the spatial arrangement of pigments, employing helices as scaffolds. In this study, we have developed porphyrin-peptoid (pigment-helix) conjugates (PPCs) that can modulate the donor-acceptor energy transfer efficiency with exceptional precision by controlling the relative distance and orientation of the two pigments. Five donor-acceptor molecular dyads were constructed using zinc porphyrin and free base porphyrin (Zn(i + 2)–Zn(i + 6)), and highly efficient energy transfer was demonstrated with estimated efficiencies ranging from 92% to 96% measured by static fluorescence emission in CH2Cl2 and from 96.3% to 97.6% using femtosecond transient absorption measurements in toluene, depending on the relative spatial arrangement of the donor-acceptor pairs. Our results suggest that the remarkable precision and tunability exhibited by nature can be achieved by mimicking the design principles of natural photosynthetic proteins.

5. Calculation of energy transfer efficiency 12 Table S2. Emission intensity at 605 nm (F605) and estimated energy transfer efficiencies of Zn PPCs.
Synthesis of porphyrin-peptoid conjugates displaying one Zn porphyrin and one free base porphyrin (Zn PPCs)

Porphyrin conjugation
Initially, the conjugation of Zn porphyrin was attempted with compound 2 using ZnTPP-NHS. ZnTPP conjugation was performed under basic conditions; however, analytical HPLC or ESI-MS showed no product.
The data indicated that ZnTPP-NHS and even compound 2 were decomposed under the reaction conditions (data not shown). Alternatively, the conjugation of Zn porphyrin with ZnTPP-COOH was attempted, without using the NHS ester ( Figure S1). The coupling agents HATU, HCTU, and COMU were tested with bases with different pKa values that are generally used in peptide coupling reactions: DIEA (N,N-diisopropylethylamine; pKa = 10.7) and NMM (N-methylmorpholine; pKa = 7.4). Unfortunately, the yields under these reaction conditions were poor; it is likely that the interactions between Zn porphyrin and coupling reagent compete with the coupling reaction, resulting in reduced efficiency. 6 At this point, we decided to avoid ZnTPP coupling to compound 2; instead, metalation of compound 2, followed by fbTPP conjugation appeared to be a feasible alternative.

Solid phase extraction (SPE) purification
Demetallation of Zn readily occurs under acidic conditions. Since we used 0.1% TFA containing buffer as HPLC mobile phases, collected fractions after preparative HPLC contained small amount of TFA. Routine practice was to lyophilize the fractions containing desired product; however, we observed release of Zn 2+ ions from porphyrin during the lyophilization, lowering purity of the final product. Therefore, SPE (solid phase extraction) was used to remove the small amount of TFA before concentrating the HPLC fractions. SPE cartridges were purchased from Waters (Oasis HLB 6 cc, 500 mg

Molecular simulations of porphyrin-peptoid conjugates (PPCs)
To gain insight into how peptoid conformational properties are affected by the spacing of porphyrin-conjugated residues, we performed ~1.2 μs replica-exchange molecular dynamics (REMD) simulations of (i, i+2) and (i, i+3) PPCs with two fbTPPs attached (or Fb(i+2) and Fb(i+3), respectively)7, using methods similar to previous approaches. 8,9,10 In these simulations, 20 or 24 temperature replicas ranging from 300 K to 800 K, with nearesttemperature exchange attempts every 10 ps, were used to efficiently sample peptoid conformations separated by large cis/trans amide barriers trajectory. As each replica performs a random walk in temperature, omega-angles of all peptoid residues decorrelate over timescales ranging from 20 to 400 ns, indicating converged sampling.
We define a peptoid residue to be helical if it has a cis-amide and a negative backbone phi angle. Per-residue

System preparation.
Molecular topologies for molecules Fb(i+2) and Fb(i+3) were constructed using the General Amber Force Field (GAFF) 11 with partial changes from AM1-BCC. 12 The GAFF+φ torsion correction was applied to all peptoid residues, as described in Mukherjee et al. 10

MD simulation. (1.2 μs × 24 (or 20) temperatures replicas from 300 K to 800 K)
The OBC Generalized Born implicit solvation model was used in all simulations. 13 Swaps between adjacent replicas were attempted every 10 ps, with acceptance ratios ranging from 0.45 to 0.51. Simulations were performed with GROMACS 4.6 dynamics package. Stochastic integration (Langevin dynamics) was used with a 9 2 fs time step and water-like viscosity. Trajectory snapshots and energies were written every 10 ps. The initial starting structure of each replica was a right-handed helical conformation with an all-cis amide backbone.
Sampling convergence was assessed using autocorrelation analysis ( Figure S5). The MDTraj package 14 was used calculate structural observables (cis-amide populations, helix populations, inter-porphyrin distances and angles) from samples after 500 ns at 300 K. Normal vectors for each porphyrin ring were calculated as the mean cross-product of sequential nitrogen-nitrogen distance vectors around the ring. As an additional test of the robustness of the results, we also performed simulations in which the GAFF+φ torsion correction was applied to all residues, including Nlys(fbTPP). Similar results were obtained in all cases ( Figure S6  Three main conformational basins can be identified on the free energy landscape for Fb(i+2) (a-c) and Fb(i+3) (d-f), each shown with a representative conformation from the simulations. Among the predicted conformations, only (a) and (d) of Fb(i+2) and Fb(i+3), respectively, show maintenance of helical peptoid conformation, which agrees with our previous experimental data (e.g., UV-vis and circular dichroism spectroscopy). 7

Calculation of energy transfer efficiency
To estimate the energy transfer efficiency of each Zn PPC, the ratio of emission intensity (i.e., F605) between Znref and Zn(i+n) was used (Table S2)

Time-resolved fluorescence of fbTPP and ZnTPP
To  Figure S8.
The emission kinetics of fbTPP and ZnTPP were fit by a convoluted model (shown in equation S1) between the Gaussian instrumental response function and an exponential decay function, where A0 is a constant, A1 is an amplitude, t0 is the position of time zero, and  is related to the pulsewidth of instrumental Gaussian function (pulsewidth = 2.325 × ), and τ is the lifetime of an exponential decay. The emission lifetimes of fbTPP and ZnTPP were determined as 12.2 ns and 2.63 ns, respectively.

Transient absorption spectroscopy
A femtosecond transient absorption spectroscopy setup based on a Ti:sapphire laser system (LIBRA-USP-HE, Coherent, Inc.; <50 fs, 1 kHz repetition) was used to measure time-resolved spectra of fbTPP, ZnTPP, and Zn PPCs. 16,17 The 547 nm pump pulses with ~35 nm FWHM were generated from a home-built non-collinear optically parametric amplifier and compressed in a prism pair compressor. A broadband probe pulses (450-1000 nm) generated from the supercontinuum generation in a sapphire window were measured in a fiber-based spectrometer (QE65Pro, Ocean Optics). 16

Global analysis
The kinetics for the excited-state dynamics and ultrafast energy transfers of transient absorption signals were analyzed by the global fit offered in the Glotaran software. 18,19 The time-resolved spectra, Ψ(t,), are analyzed as a superposition of the specific kinetic components as shown in equation S2, 1 ( , ) ( ) ( )