Mingzhen Zhang

Engineering antimicrobial peptides with improved antimicrobial and hemolytic activities.

The rapid rise of antibiotic resistance in pathogens becomes a serious and growing threat to medicine and public health. Naturally occurring antimicrobial peptides (AMPs) are an important line of defense in the immune system against invading bacteria and microbial infection. In this work, we present a combined computational and experimental study of the biological activity and membrane interaction of the computationally designed Bac2A-based peptide library. We used the MARTINI coarse-grained molecular dynamics with adaptive biasing force method and the umbrella sampling technique to investigate the translocation of a total of 91 peptides with different amino acid substitutions through a mixed anionic POPE/POPG (3:1) bilayer and a neutral POPC bilayer, which mimic the bacterial inner membrane and the human red blood cell (hRBC) membrane, respectively. Potential of mean force (PMF, free energy profile) was obtained to measure the free energy barrier required to transfer the peptides from the bulk water phase to the water-membrane interface and to the bilayer interior. Different PMF profiles can indeed identify different membrane insertion scenarios by mapping out peptide-lipid energy landscapes, which are correlated with antimicrobial activity and hemolytic activity. Computationally designed peptides were further tested experimentally for their antimicrobial and hemolytic activities using bacteria growth inhibition assay and hemolysis assay. Comparison of PMF data with cell assay results reveals a good correlation of the peptides between predictive transmembrane activity and antimicrobial/hemolytic activity. Moreover, the most active mutants with the balanced substitutions of positively charged Arg and hydrophobic Trp residues at specific positions were discovered to achieve the improved antimicrobial activity while minimizing red blood cell lysis. Such substitutions provide more effective and cooperative interactions to distinguish the peptide interaction with different lipid bilayers. This work provides a useful computational tool to better understand the mechanism and energetics of membrane insertion of AMPs and to rationally design more effective AMPs.

Synthesis and Characterization of Antifouling Poly(N-acryloylaminoethoxyethanol) with Ultralow Protein Adsorption and Cell Attachment

Rational design of effective antifouling polymers is challenging but important for many fundamental and applied applications. Herein we synthesize and characterize an N-acryloylaminoethoxyethanol (AAEE) monomer, which integrates three hydrophilic groups of hydroxyl, amide, and ethylene glycol in the same material. AAEE monomers were further grafted and polymerized on gold substrates to form polyAAEE brushes with well-controlled thickness via surface-initiated atomic transfer radical polymerization (SI-ATRP), with particular attention to a better understanding of the molecular structure-antifouling property relationship of hydroxyl-acrylic-based polymers. The surface hydrophilicity and antifouling properties of polyAAEE brushes as a function of film thickness are studied by combined experimental and computational methods including surface plasmon resonance (SPR) sensors, atomic force microscopy (AFM), cell adhesion assay, and molecular dynamics (MD) simulations. With the optimal polymer film thicknesses (∼10-40 nm), polyAAEE-grafted surfaces can effectively resist protein adsorption from single-protein solutions and undiluted human blood plasma and serum to a nonfouling level (i.e., <0.3 ng/cm(2)). The polyAAEE brushes also highly resist mammalian cell attachment up to 3 days. MD simulations confirm that the integration of three hydrophilic groups induce a stronger and closer hydration layer around polyAAEE, revealing a positive relationship between surface hydration and antifouling properties. The molecular structure-antifouling properties relationship of a series of hydroxyl-acrylic-based polymers is also discussed. This work hopefully provides a promising structural motif for the design of new effective antifouling materials beyond traditional ethylene glycol-based antifouling materials.

Probing the Structural Dependence of Carbon Space Lengths of Poly(N-hydroxyalkyl acrylamide)-Based Brushes on Antifouling Performance

Numerous biocompatible antifouling polymers have been developed for a wide variety of fundamental and practical applications in drug delivery, biosensors, marine coatings, and many other areas. Several antifouling mechanisms have been proposed, but the exact relationship among molecular structure, surface hydration property, and antifouling performance of antifouling polymers still remains elusive. Here this work strives to provide a better understanding of the structure-property relationship of poly(N-hydroxyalkyl acrylamide)-based materials. We have designed, synthesized, and characterized a series of polyHAAA brushes of various carbon spacer lengths (CSLs), that is, poly(N-hydroxymethyl acrylamide) (polyHMAA), poly(N-(2-hydroxyethyl)acrylamide) (polyHEAA), poly(N-(3-hydroxypropyl)acrylamide) (polyHPAA), and poly(N-(5-hydroxypentyl)acrylamide) (polyHPenAA), to study the structural dependence of CSLs on their antifouling performance. HMAA, HEAA, HPAA, and HPenAA monomers contained one, two, three, and five methylene groups between hydroxyl and amide groups, while the other groups in polymer backbones were the same as each other. The relation of such small structural differences of polymer brushes to their surface hydration and antifouling performance was studied by combined experimental and computational methods including surface plasmon resonance sensors, sum frequency generation (SFG) vibrational spectroscopy, cell adhesion assay, and molecular simulations. Antifouling results showed that all polyHAAA-based brushes were highly surface resistant to protein adsorption from single protein solutions, undiluted blood serum and plasma, as well as cell adhesion up to 7 days. In particular, polyHMAA and polyHEAA with the shorter CSLs exhibited higher surface hydration and better antifouling ability than polyHPMA and polyHPenAA. SFG and molecular simulations further revealed that the variation of CSLs changed the ratio of hydrophobicity/hydrophilicity of polymers, resulting in different hydration characteristics. Among them, polyHMAA and polyHEAA with the shorter CSLs showed the highest potency for surface hydration and antifouling abilities, while polyHPenAA showed the lowest potency. The combination of both hydroxyl and amide groups in the same polymer chain provides a promising structural motif for the design of new effective antifouling materials.

Cross-Seeding Interaction between β-Amyloid and Human Islet Amyloid Polypeptide.

Alzheimers disease (AD) and type 2 diabetes (T2D) are two common protein misfolding diseases. Increasing evidence suggests that these two diseases may be correlated with each other via cross-sequence interactions between β-amyloid peptide (Aβ) associated with AD and human islet amyloid polypeptide (hIAPP) associated with T2D. However, little is known about how these two peptides work and how they interact with each other to induce amyloidogenesis. In this work, we study the effect of cross-sequence interactions between Aβ and hIAPP peptides on hybrid amyloid structures, conformational changes, and aggregation kinetics using combined experimental and simulation approaches. Experimental results confirm that Aβ and hIAPP can interact with each other to aggregate into hybrid amyloid fibrils containing β-sheet-rich structures morphologically similar to pure Aβ and hIAPP. The cross-seeding of Aβ and hIAPP leads to the coexistence of both a retarded process at the initial nucleation stage and an accelerated process at the fibrillization stage, in conjunction with a conformational transition from random structures to α-helix to β-sheet. Further molecular dynamics simulations reveal that Aβ and hIAPP oligomers can efficiently cross-seed each other via the association of two highly similar U-shaped β-sheet structures; thus, conformational compatibility between Aβ and hIAPP aggregates appears to play a key role in determining barriers to cross-seeding. The cross-seeding effects in this work may provide new insights into the molecular mechanisms of interactions between AD and T2D.

Structural and Energetic Insight into the Cross-Seeding Amyloid Assemblies of Human IAPP and Rat IAPP

The misfolding and aggregation of human islet amyloid polypeptide (hIAPP or amylin) into small oligomers and large amyloid fibrils is believed to be responsible for the dysfunction and death of pancreatic β-cells in diabetes type II. However, rat IAPP (rIAPP), which differs from the hIAPP by only 6 of 37 residues, lacks the ability to form amyloid fibrils and to induce cell death. Little is known about the cross-sequence interactions and cross-seeding structures between hIAPP and rIAPP peptides. Herein using explicit-solvent molecular dynamics (MD) simulations, we modeled and simulated different heteroassemblies formed by the amyloidogenic hIAPP and the nonamyloidogenic rIAPP peptides. Simulations showed that the U-shaped hIAPP monomer and oligomers can interact with conformationally similar rIAPP to form stable complexes and to coassemble into heterogeneous structures. Stable heterointeractions between hIAPP and rIAPP were shown to arise from hydrophobic contacts and hydrogen bonds at the interface, particularly at N- and C-terminal β-sheet regions. Because of the enhanced interpeptide interactions at the interface, upon binding to hIAPP oligomers, the β-sheet population of rIAPP was greatly increased as compared to that of rIAPP alone. More importantly, the conformational energies of rIAPP monomers at the bound state were observed to be consistently higher than those of rIAPP monomers at the unbound state. However, rIAPP monomers enable one to adopt different conformations and follow different pathways for associating with hIAPP from the high energy of the bound state to the low energy of the unbound state, without encountering any large and abrupt energy barrier. In parallel, AFM study of cross-aggregation of hIAPP and rIAPP provided additional evidence that hIAPP can seed with rIAPP to form hybrid fibrils at all concentrations similar to pure hIAPP fibrils. This work demonstrates the existence of cross-interactions between the two different IAPP peptides, which provides an improved fundamental understanding of the cross-seeding of different amyloid sequences toward amyloid aggregation and toxicity mechanisms.

Molecular Understanding and Structural-Based Design of Polyacrylamides and Polyacrylates as Antifouling Materials

Design and synthesis of highly bioinert and biocompatible antifouling materials are crucial for a broad range of biomedical and engineering applications. Among antifouling materials, polyacrylamides and polyacrylates have proved so promising because of cheap raw materials, ease of synthesis and applicability, and abundant functional groups. The strong surface hydration and the high surface packing density of polyacrylamides and polyacrylates are considered to be the key contributors to their antifouling property. In this article, we review our studies on the design and synthesis of a series of polyacrylamides and polyacrylates with different molecular structures. These polymers can be fabricated into different architectural forms (brushes, nanoparticles, nanogels, and hydrogels), all of which are highly resistant to the attachment of proteins, cells, and bacteria. We find that small structural changes in the polymers can lead to large enhancement in surface hydration and antifouling performance, both showing a positive correlation. This reveals a general design rule for effective antifouling materials. Furthermore, polyacrylamides and polyacrylates are readily functionalized with other bioactive compounds to achieve different new multifunctionalities.

Cross-Sequence Interactions between Human and Rat Islet Amyloid Polypeptides

Human islet amyloid polypeptide (hIAPP) can assemble into toxic oligomers and fibrils, which are associated with cell degeneration and the pathogenesis of type 2 diabetes. Cross-interaction of hIAPP with rat IAPP (rIAPP)--a non-amyloidogenic peptide with high sequence similarity to hIAPP--might influence the aggregation and toxicity of hIAPP. However, the exact role of rIAPP in hIAPP aggregation and toxicity still remains unclear. In this work, we investigated the effect of cross-sequence interactions between full-length hIAPP(1-37) and rIAPP(1-37) on hybrid amyloid structures, aggregation kinetics, and cell toxicity using combined computational and experimental approaches. Experimental results indicate a contrasting role of rIAPP in hIAPP aggregation, in which rIAPP initially inhibits the early aggregation and nuclei formation of hIAPP, but hIAPP seeds can also recruit both hIAPP and rIAPP to form more hybrid fibrils, thus promoting amyloid fibrillation ultimately. The coincubation of hIAPP and rIAPP also decreases cell viability, presumably due to the formation of more toxic hybrid oligomers at the prolonged lag phase. Comparative MD simulations confirm that the cross-sequence interactions between hIAPP and rIAPP stabilize β-sheet structure and thus likely promote their fibrillization. This work provides valuable insights into a critical role of cross-amyloid interactions in protein aggregation.

Polymorphic Associations and Structures of the Cross-Seeding of Aβ1–42 and hIAPP1–37 Polypeptides

Emerging evidence have shown that the patients with Alzheimers disease (AD) often have a higher risk of later developing type II diabetes (T2D), and vice versa, suggesting a potential pathological link between AD and T2D. Amyloid-β (Aβ) and human islet amyloid polypeptide (hIAPP) are the principle causative components responsible for the pathologies of AD and T2D, respectively. The cross-sequence interactions between Aβ and hIAPP may provide a molecular basis for better understanding the potential link between AD and T2D. Herein, we systematically modeled and simulated the cross-sequence aggregation process, molecular interactions, and polymorphic structures of full-length Aβ and hIAPP peptides using a combination of coarse-grained (CG) replica-exchange molecular dynamics (REMD) and all-atom molecular dynamics (MD) simulations, with particular focus on the effect of association models between Aβ and hIAPP on the structural stability and polymorphic populations of hybrid Aβ-hIAPP aggregates. Four distinct association models (double-layer, elongation, tail-tail, and block models) between Aβ and hIAPP oligomers were identified, and the associated polymorphic Aβ-hIAPP structures were determined as well. Among them, different association models led to different Aβ-hIAPP aggregates, with large differences in structural morphologies and populations, interacting interfaces, and underlying association forces. The computational models support the cross-sequence interactions between Aβ and hIAPP pentamers, which would lead to the complex hybrid Aβ-hIAPP assemblies. This computational work may also provide a different point of view to a better understanding of a potential link between AD and T2D.

Membrane Interactions of hIAPP Monomer and Oligomer with Lipid Membranes by Molecular Dynamics Simulations

Interaction of human islet amyloid polypeptide (hIAPP) peptides with cell membrane is crucial for the understanding of amyloid toxicity associated with Type II diabetes (T2D). While it is known that the hIAPP-membrane interactions are considered to promote hIAPP aggregation into fibrils and induce membrane disruption, the membrane-induced conformation, orientation, aggregation, and adsorption behaviors of hIAPP peptides have not been well understood at the atomic level. Herein, we perform all-atom explicit-water molecular dynamics (MD) simulations to study the adsorption, orientation, and surface interaction of hIAPP aggregates with different sizes (monomer to tetramer) and conformations (monomer with α-helix and tetramer with β-sheet-rich U-turn) upon adsorption on the lipid bilayers composed of both pure zwitterionic POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) and mixed anionic POPC/POPE (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine) (3:1) lipids. MD simulation results show that hIAPP monomer with α-helical conformation and hIAPP pentamer with β-sheet conformation can adsorb on both POPC and POPC/POPE bilayers via a preferential orientation of N-terminal residues facing toward the bilayer surface. The hIAPP aggregates show stronger interactions with mixed POPC/POPE lipids than pure POPC lipids, consistent with experimental observation that hIAPP adsorption and fibrililation are enhanced on mixed lipid bilayers. While electrostatic interactions are main attractive forces to drive the hIAPP aggregates to adsorb on both bilayers, the introduction of the more hydrophilic head groups of POPE lipids further promote the formation of the interfacial hydrogen bonds. Complement to our previous studies of hIAPP aggregates in bulk solution, this computational work increases our knowledge about the mechanism of amyloid peptide-membrane interactions that is central to the understanding the progression of all amyloid diseases.

Comparative Study of Graphene Hydrogels and Aerogels Reveals the Important Role of Buried Water in Pollutant Adsorption

Water as the universal solvent has well-demonstrated its ability to dissolve many substances, but buried water inside different nanoporous materials always exhibits some unusual behaviors. Herein, 3D porous graphene hydrogel (GH) is developed as a super-adsorbent to remove different pollutants (antibiotics, dyes, and heavy ions) for water purification. Due to its highly porous structure and high content of water, GH also demonstrated its super adsorption capacity for adsorbing and removing different pollutants (antibiotics, dyes, and heavy ions) as compared to conventional graphene aerogel (GA). More fundamentally, the buried-water enhanced adsorption mechanism was proposed and demonstrated, such that buried water in GH plays the combinatorial roles as (1) supporting media, (2) transport nanochannels, and (3) hydrogen bondings in promoting pollutant adsorption. In parallel, molecular dynamics simulations further confirm that buried water in GH has the stronger interaction with pollutants via hydrogen bonds than other buried alcohols. GH integrates the merit of both graphene (e.g., fine chemical resistance and excellent mechanical property) and hydrogel (e.g., high water content, porous structure, and simple solution-based processability and scalability), giving it promising potential for environmental applications.