Biological fluids, the most complex blends, have compositions that constantly vary and cannot be molecularly defined1. Despite these uncertainties, proteins fluctuate, fold, function and evolve as programmed2,3,4. We propose that in addition to the known monomeric sequence requirements, protein sequences encode multi-pair interactions at the segmental level to navigate random encounters5,6; synthetic heteropolymers capable of emulating such interactions can replicate how proteins behave in biological fluids individually and collectively. Here, we extracted the chemical characteristics and sequential arrangement along a protein chain at the segmental level from natural protein libraries and used the information to design heteropolymer ensembles as mixtures of disordered, partially folded and folded proteins. For each heteropolymer ensemble, the level of segmental similarity to that of natural proteins determines its ability to replicate many functions of biological fluids including assisting protein folding during translation, preserving the viability of fetal bovine serum without refrigeration, enhancing the thermal stability of proteins and behaving like synthetic cytosol under biologically relevant conditions. Molecular studies further translated protein sequence information at the segmental level into intermolecular interactions with a defined range, degree of diversity and temporal and spatial availability. This framework provides valuable guiding principles to synthetically realize protein properties, engineer bio/abiotic hybrid materials and, ultimately, realize matter-to-life transformations.
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All data needed to evaluate the conclusions in the paper are present in the paper and/or the supplementary materials. For reproduction purposes, the raw data used to generate the figures are available from the Dryad Digital Repository (DOI:10.6078/D1KH8R ).
Source code and input scripts supporting this work are available at https://github.com/Shunili/AE-RHP.
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The work was supported by the US Department of Defense (DOD), Army Research Office, under contract no. W911NF-13-1-0232, Defense Threat Reduction Agency (DTRA) under contract no. HDTRA1-19-1-0011, the National Science Foundation under contract no. DMR-2104443, the US Department of Energy, Office of Science, Office of Basic Energy Sciences, Materials Sciences and Engineering Division, under contract no. DE-AC02-05-CH11231 (KC3104) and the Alfred P. Sloan Foundation (grant no. G-2021-16757). Z.R. is supported by the Kavli Energy NanoScience Institute through the Kavli ENSI Philomathia Graduate Student Fellowship Program. Scattering studies were done at Advanced Photon Source and use of the Advanced Photon Source was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, under contract no. DE-AC02-06CH1135.
T.X., H.H., Z.R. and S.L. have a pending PCT patent application. The rest of the authors declare no competing interests.
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Extended data figures and tables
Extended Data Fig. 1 Blocks and 50-mers along a polymer chain.
(a) Each monomer was reassigned to one of two pseudo-monomers (hydrophobic vs. hydrophilic) based on monomer’s hydrophobicity. A block comprises consecutive monomers of a single type. (b) A polymer chain was truncated into a set of 50-mers.
Extended Data Fig. 2 The FBS solution with different RHP ensembles after incubating at 52 °C for 2.5 h.
The FBS solution forms a thin film (like “milk skin”) at the air-water interface after thermal treatment (red circle). RHP4 exhibits the weakest tendency for formation of this thin film among three tested RHP ensembles. The solution is stirred to tear the thin film into suspended flakes for visualization.
Extended Data Fig. 3 Temperature-dependent 1H-NMR spectrum of RHP4 in D2O.
(a) 1H-NMR spectrum of RHP4 in D2O at 37 °C and its chemical structure. (b) The FWHM of proton peaks (a, p, and n) as a function of temperature (30–70 °C). (c) Part of 1H-NMR spectra of RHP4 as a function of temperature (30–70 °C) including proton peaks of the OEGMA side chain.
Extended Data Fig. 4 The effect of solvent polarity on 1H-NMR spectrum of RHP4 in d6-DMSO/D2O cosolvent
Extended Data Fig. 5 The distribution of DHP2 segments in PCA space.
From top to bottom, the distribution of segments from hydrophobic region, amphiphilic region, and hydrophilic region along DHP2 chains were shown.
Extended Data Fig. 6 Differential interference contrast (DIC) images of (a) RHP8 (b) RHP9 phase-separated droplets.
Each sample is 1 mg/ml in sodium phosphate buffer (50 mM, pH 7.0).
Extended Data Fig. 7 Folding status of AqpZ-eGFP in the presence of 0.2 wt% RHPs based on the eGFP fluorescence.
Error bar is 1 s.d and n = 3.
Extended Data Fig. 8 Temperature-dependent turbidimetry for RHP14 ensemble.
The polymer solution is 1mg/ml in sodium phosphate buffer (50 mM, pH 7.0).
Extended Data Fig. 9 FRAP analysis of liquid-like coacervates made from RHP10 ensemble.
The recovery trace shows the normalized recovery of a bleached region. Solid red curve fits to an exponential function (see FRAP method section).
This file contains Supplementary Tables 1 and 2 and Figs. 1–21.
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Ruan, Z., Li, S., Grigoropoulos, A. et al. Population-based heteropolymer design to mimic protein mixtures. Nature 615, 251–258 (2023). https://doi.org/10.1038/s41586-022-05675-0
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