Pathway-level acceleration of glycogen catabolism by a response regulator in the cyanobacterium Synechocystis species PCC 6803.

Response regulators of two-component systems play pivotal roles in the transcriptional regulation of responses to environmental signals in bacteria. Rre37, an OmpR-type response regulator, is induced by nitrogen depletion in the unicellular cyanobacterium Synechocystis species PCC 6803. Microarray and quantitative real-time polymerase chain reaction analyses revealed that genes related to sugar catabolism and nitrogen metabolism were up-regulated by rre37 overexpression. Protein levels of GlgP(slr1367), one of the two glycogen phosphorylases, in the rre37-overexpressing strain were higher than those of the parental wild-type strain under both nitrogen-replete and nitrogen-depleted conditions. Glycogen amounts decreased to less than one-tenth by rre37 overexpression under nitrogen-replete conditions. Metabolome analysis revealed that metabolites of the sugar catabolic pathway and amino acids were altered in the rre37-overexpressing strain after nitrogen depletion. These results demonstrate that Rre37 is a pathway-level regulator that activates the metabolic flow from glycogen to polyhydroxybutyrate and the hybrid tricarboxylic acid and ornithine cycle, unraveling the mechanism of the transcriptional regulation of primary metabolism in this unicellular cyanobacterium.

Increased bioplastic production with an RNA polymerase sigma factor SigE during nitrogen starvation in Synechocystis sp. PCC 6803.

Because cyanobacteria directly harvest CO2 and light energy, their carbon metabolism is important for both basic and applied sciences. Here, we show that overexpression of the sigma factor sigE in Synechocystis sp. PCC 6803 widely changes sugar catabolism and increases production of the biodegradable polyester polyhydroxybutyrate (PHB) during nitrogen starvation. sigE overexpression elevates the levels of proteins implicated in glycogen catabolism, the oxidative pentose phosphate pathway, and polyhydroxyalkanoate biosynthesis. PHB accumulation is enhanced by sigE overexpression under nitrogen-limited conditions, yet the molecular weights of PHBs synthesized by the parental glucose-tolerant and sigE overexpression strain are similar. Although gene expression induced by nitrogen starvation is changed and other metabolites (such as GDP-mannose and citrate) accumulate under sigE overexpression, genetic engineering of this sigma factor altered the metabolic pathway from glycogen to PHB during nitrogen starvation.

Characterization of site-specific mutations in a short-chain-length/medium-chain-length polyhydroxyalkanoate synthase: in vivo and in vitro studies of enzymatic activity and substrate specificity.

Saturation point mutagenesis was carried out at position 479 in the polyhydroxyalkanoate (PHA) synthase from Chromobacterium sp. strain USM2 (PhaC(Cs)) with specificities for short-chain-length (SCL) [(R)-3-hydroxybutyrate (3HB) and (R)-3-hydroxyvalerate (3HV)] and medium-chain-length (MCL) [(R)-3-hydroxyhexanoate (3HHx)] monomers in an effort to enhance the specificity of the enzyme for 3HHx. A maximum 4-fold increase in 3HHx incorporation and a 1.6-fold increase in PHA biosynthesis, more than the wild-type synthase, was achieved using selected mutant synthases. These increases were subsequently correlated with improved synthase activity and increased preference of PhaC(Cs) for 3HHx monomers. We found that substitutions with uncharged residues were beneficial, as they resulted in enhanced PHA production and/or 3HHx incorporation. Further analysis led to postulations that the size and geometry of the substrate-binding pocket are determinants of PHA accumulation, 3HHx fraction, and chain length specificity. In vitro activities for polymerization of 3HV and 3HHx monomers were consistent with in vivo substrate specificities. Ultimately, the preference shown by wild-type and mutant synthases for either SCL (C(4) and C(5)) or MCL (C(6)) substrates substantiates the fundamental classification of PHA synthases.

Monitoring and kinetic analysis of the molecular interactions by which a repressor protein, PhaR, binds to target DNAs and poly[(R)-3-hydroxybutyrate].

The repressor protein PhaR, which is a component of poly[(R)-3-hydroxybutyrate] granules, functions as a repressor of the gene expression of the phasin PhaP and of PhaR itself. We used a quartz crystal microbalance to investigate the binding behavior by which PhaR in Ralstonia eutropha H16 targets DNAs and amorphous poly[(R)-3-hydroxybutyrate] thin films. Binding rate constants, dissociation rate constants, and dissociation constants of the binding of PhaR to DNA and to amorphous poly[(R)-3-hydroxybutyrate] suggested that PhaR bind to both in a similar manner. On the basis of the binding rate constant values, we proposed that the phaP gene would be derepressed in harmony with the ratio of the concentration of the target DNA to the concentration of amorphous poly[(R)-3-hydroxybutyrate] at the start of poly[(R)-3-hydroxybutyrate] synthesis in R. eutropha H16.

Active intermediates of polyhydroxyalkanoate synthase from Aeromonas caviae in polymerization reaction.

Polyhydroxyalkanoate (PHA) synthase from Aeromonas caviae FA440 (PhaC(Ac), BAA21815) is one of the most valuable PHA synthase, because of its function to synthesize a practical bioplastic, poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)]. However, biochemical activity and active intermediates of PhaC(Ac) have not been clarified until now. In the present study, a gene of PhaC(Ac) was cloned and overexpressed by a cell-free protein expression system. Both the polymerization activity and oligomerization behavior of the purified PhaC(Ac) were characterized in order to clarify the active intermediates of PhaC(Ac) based on the hydrodynamic diameters and specific activities of PhaC(Ac). The influences of a substrate, (R)-3-hydroxybutyryl-CoA (3HB-CoA), on the oligomerization of PhaC(Ac) (7.5 µM) were also investigated, and then the Hill coefficient (n = 2.6 ± 0.4) and the microscopic dissociation constant (K(m) = 77 ± 5 µM) were determined. Based on the results, the active intermediate of PhaC(Ac) was concluded to be the dimeric PhaC(Ac) containing 3HB-CoA as an activator for its dimerization. This information is critical for revealing the relationships between its dimerization and function in PHA synthesis.

Biosynthesis of polyhydroxyalkanaotes by a novel facultatively anaerobic Vibrio sp. under marine conditions.

Marine bacteria have recently attracted attention as potentially useful candidates for the production of practical materials from marine ecosystems, including the oceanic carbon dioxide cycle. The advantages of using marine bacteria for the biosynthesis of poly(hydroxyalkanoate) (PHA), one of the eco-friendly bioplastics, include avoiding contamination with bacteria that lack salt-water resistance, ability to use filtered seawater as a culture medium, and the potential for extracellular production of PHA, all of which would contribute to large-scale industrial production of PHA. A novel marine bacterium, Vibrio sp. strain KN01, was isolated and characterized in PHA productivity using various carbon sources under aerobic and aerobic-anaerobic marine conditions. The PHA contents of all the samples under the aerobic-anaerobic condition, especially when using soybean oil as the sole carbon source, were enhanced by limiting the amount of dissolved oxygen. The PHA accumulated using soybean oil as a sole carbon source under the aerobic-anaerobic condition contained 14% 3-hydroxypropionate (3HP) and 3% 5-hydroxyvalerate (5HV) units in addition to (R)-3-hydroxybutyrate (3HB) units and had a molecular weight of 42 × 10³ g/mol. The present result indicates that the activity of the beta-oxidation pathway under the aerobic-anaerobic condition is reduced due to a reduction in the amount of dissolved oxygen. These findings have potential for use in controlling the biosynthesis of long main-chain PHA by regulating the activity of the beta-oxidation pathway, which also could be regulated by varying the dissolved oxygen concentration.

Effect of phase structure on enzymatic degradation in poly(L-lactide)/atactic poly(3-hydroxybutyrate) blends with different miscibility.

Thin films of poly(L-lactide) (PLLA)/atactic poly(3-hydroxybutyrate) (ataPHB) blends with different miscibility were prepared and characterized by using differential scanning calorimetry (DSC) and atomic force microscopy (AFM). The DSC analysis suggested that the blend thin films exhibited different phase structures, such as miscible, partially miscible, and immiscible depending on the blending ratio as well as molecular weight of ataPHB component. The different miscibility was further confirmed by the surface morphological observation by AFM. Both the immiscible and partially miscible blends of PLLA/ataPHB revealed the formation of phase-separated morphology of PLLA and ataPHB components, whereas the homogeneous surface morphology was observed for the miscible blend. On the basis of the changes in the depth profile from the surface level of the thin films, the enzymatic degradation rates of the PLLA and ataPHB domains were determined in the presence of either PHB depolymerase or proteinase K, respectively. The erosion rate of PLLA/ataPHB blends was strongly dependent on the blend composition and the degree of dispersion of the two components. The enzymatic degradation behaviors were discussed in terms of phase structure, molecular mobility, and retardation effect of the components in the blends.

Chimeric enzyme composed of polyhydroxyalkanoate (PHA) synthases from Ralstonia eutropha and Aeromonas caviae enhances production of PHAs in recombinant Escherichia coli.

Chimeric enzymes composed of polyhydroxyalkanoate (PHA) synthases from Ralstonia eutropha (Cupriavidus necator) (PhaC(Re)) and Aeromonas caviae (PhaC(Ac)) were constructed. PhaC(Re) is known for its potent enzymatic activity among the characterized PHA synthases. PhaCAc has broad substrate specificity and synthesizes short-chain-length (SCL)/medium-chain-length (MCL) PHA. We attempted to create chimeric enzymes inheriting both of the advantageous properties. Among eight chimeras, AcRe12, with 26% of the N-terminal of PhaC(Ac) and 74% of the C-terminal of PhaC(Re), exhibited comparable P(3-hydroxybutyrate) accumulation as parental enzymes in Escherichia coli JM109. Thus, AcRe12 was applied to SCL/MCL PHA production using E. coli LS5218 as the host. AcRe12 accumulated higher amount of PHA (50 wt %) than the parental enzymes. Furthermore, the PHA consisted of 2 mol % 3-hydroxyhexanoate as well as 3-hydroxybutyrate. Therefore, the chimeric PHA synthase, AcRe12, inherited the character of both of the parental enzymes and thus exhibits improved enzymatic properties.

Evaluating the ability of polyhydroxyalkanoate synthase mutants to produce P(3HB-co-3HA) from soybean oil.

Polyhydroxyalkanoate (PHA) synthase from Pseudomonas sp 61-3 (PhaC1(Ps)) is able to synthesize P(3HB-co-3HA), consisting of a 3HB unit and medium-chain-length 3HA units of 6-12 carbon atoms. Expression vectors encoding 76 PhaC1(Ps) mutants with an amino acid replacement at position 130, 325, 477 or 481 were individually introduced into Ralstonia eutropha. The mutant enzyme genes were evaluated in terms of their abilities to synthesize P(3HB-co-3HA) using soybean oil as a carbon source. 20 mutants showed significantly high accumulation levels of PHA exceeding 30 wt.-% and as high as 57 wt.-%. It was found that hydrophobic amino acids at the positions are more likely to enhance accumulation of PHA in R. eutropha.

Cloning of poly(aspartic acid) (PAA) hydrolase-1 gene from Pedobacter sp. KP-2 and hydrolysis of thermally synthesized PAA by its gene product.

Pedobacter sp. KP-2 can degrade and metabolize thermally synthesized alpha,beta-poly(D,L-aspartic acid) (tPAA), which contains 70% of unnatural beta-amide units, with high-molecular-weight. In this study, gene cloning and molecular characterization of PAA hydrolase-1 from KP-2 was carried out. Gene analysis reveals that deduced amino acid sequence of the enzyme shows a similarity to only that of PAA hydrolase-1 from Sphingomonas sp. KT-1. GPC and NMR analyses of the hydrolyzed products of tPAA by PAA hydrolase-1 of KP-2 indicate that this enzyme cleaves the beta-beta amide linkage via endo-mode to yield oligo(aspartic acid) from tPAA. Taking the composition of tPAA and the substrate specificity of PAA hydrolase-1 into consideration, the enzyme possibly plays a crucial role in tPAA biodegradation by KP-2.

Polyhydroxyalkanoate film formation and synthase activity during in vitro and in situ polymerization on hydrophobic surfaces.

In vitro and in situ enzymatic polymerization of polyhydroxyalkanoate (PHA) on two hydrophobic surfaces, a highly oriented pyrolytic graphite (HOPG) and an alkanethiol self-assembled monolayer (SAM), was studied by atomic force microscopy (AFM) and quartz crystal microbalance (QCM), using purified Ralstonia eutropha PHA synthase (PhaC(Re)) as a biocatalyst. (R)-Specific enoyl-CoA hydratase was used to prepare R-enantiomer monomers [(R)-3-hydroxyacyl-CoA] with an acyl chain length of 4-6 carbon atoms. PHA homopolymers with different side-chain lengths, poly[(R)-3-hydroxybutyrate] [P(3HB)] and poly[(R)-3-hydroxyvalerate] [P(3HV)] were successfully synthesized from such R-enantiomer monomers on HOPG substrates. After the reaction, the surface morphologies were analyzed by AFM, revealing a nanometer thick PHA film. The same biochemical polymerization process was observed on an alkanethiol (C18) SAM surface fabricated on a gold electrode using QCM. This analysis showed that a complex sequence of PhaC(Re) adsorption and PHA polymerization has occurred on the hydrophobic surface. On the basis of these observations, the possible mechanisms of the PhaC(Re)-catalyzed polymerization reaction on the surface of hydrophobic substrates are proposed.

Enzymatic degradation of monolayer for poly(lactide) revealed by real-time atomic force microscopy: effects of stereochemical structure, molecular weight, and molecular branches on hydrolysis rates.

The influences of the stereochemical structure, the molecular weight, and the number of molecular branches for poly(lactide) (PLA) on enzymatic hydrolysis rates of PLA monolayers were studied by atomic force microscopy (AFM) and the Langmuir-Blodgett (LB) technique. Monolayers of six kinds of PLA with different molecular weights, stereochemical structure, and numbers of molecular branches were prepared by LB techniques and then characterized by AFM in air. The PLA molecules covered homogeneously with a silicon substrate and did not form lamellar crystals in the monolayer. We determined the initial hydrolysis rate of PLA monolayers in presence of proteinase K by volumetric analysis from the continuous AFM height images. The presence of D-lactyl unit reduced the hydrolysis rate of the monolayer. The hydrolysis rate for the linear PLLA samples increased with a decrease in the molecular weight. In contrast, the rates of erosion for branched PLLA monolayers were independent of the molecular weight of samples. The erosion rate of branched PLLA monolayers was found to be dependent on the average molecular weight of PLLA segment in branched molecules, not on the overall molecular weight of samples. From these results, furthermore, the hydrolysis mode of PLAs by proteinase K is discussed.

Structural effects of terminal groups on nonenzymatic and enzymatic degradations of end-capped poly(L-lactide).

Poly(L-lactide) (PLLA) with various alkyl ester chain end groups were synthesized by ring-opening polymerization of L-lactide in the presence of zinc alkoxide as a catalyst. The structural effect of chain end groups on the rate of enzymatic and nonenzymatic degradations for amorphous films of PLLA were investigated at 37 degrees C in a Tris-HCl buffer solution (pH 8.6) with proteinase K and at 60 degrees C in a phosphate buffer solution (pH 7.4), respectively. The rate of enzymatic degradation for PLLA films was dependent on the carbon numbers of alkyl ester chain end groups, and the rates of PLLA samples with dodecyl (C12), tridecyl (C13), and tetracocyl (C14) ester end groups were much lower than those of the other samples. The surface morphologies of PLLA films after enzymatic degradation were characterized by scanning electron microscopy. After the enzymatic degradation, non-end-capped PLLA, PLLA with methyl (C1) and hexyl (C6) ester chain ends, were degraded homogeneously by proteinase K and the film surface was very smooth. In contrast, the PLLA with alkyl ester chain ends of carbon numbers over 12 were degraded heterogeneously by the enzyme, and the sponge-like network structure was formed on the film surface. These results indicated that the long alkyl ester groups at the chain ends of PLLA molecules aggregated in the amorphous films and the erosion rate was depressed due to the coverage of the aggregated terminal groups on the film surface. For the nonenzymatic degradation, the molecular weight of non-end-capped PLLA was remarkably decreased with progress of degradation. In contrast, the molecular weight of the end-capped PLLA gradually reduced at the initial stage of degradation and then the rate of degradation was accelerated. The decreases of molecular weight of PLLA by autocatalyzed degradation were retarded by the capping of carboxyl chain ends.

FabG mediates polyhydroxyalkanoate production from both related and nonrelated carbon sources in recombinant Escherichia coli LS5218.

Polyhydroxyalkanoates (PHAs) composed of a mixture of short-chain-length-medium-chain-length (SCL-MCL) hydroxyacyl monomers are biologically produced polyesters that have properties ranging from thermoplastic to elastomeric, dependent on the molar ratio of SCL to MCL monomers incorporated into the copolymer. Because of the potential wide range of properties and applications for SCL-MCL PHA copolymers, it is important to develop and characterize novel metabolic pathways for SCL-MCL PHA production. The current study shows that coexpression of fabG genes from either E. coli or Pseudomonas sp. 61-3 with fabH(F87T) and PHA synthase genes enhances the production of SCL-MCL PHA copolymer from both related and nonrelated carbon sources in Escherichia coli LS5218, indicating the flexibility of FabG as a monomer-supplying enzyme for biological PHA production.

Characterization of short-chain poly3-hydroxybutyrate in baker's yeast.

A short-chain poly3-hydroxybutyrate including four comonomers, originating from a complex with calcium polyphosphate, was isolated from commercial baker's yeast cells (Saccharomyces cerevisiae) and characterized as the second complexed poly(3-hydroxyalkanoate) (cPHA) in eukaryotes. The number-average molecular weight of 4982.5 Da with a polydispersity index of 1.11 was much lower than that of beet cPHA previously isolated. End-group analysis suggested that at least 60% of the molecules form the cyclic structures. Here, the organism-dependent structural diversity of cPHAs was completely established. It was also found that a change of culture medium influences the molecular weight but not the polydispersity of baker's yeast cPHA.

Combination of N149S and D171G mutations in Aeromonas caviae polyhydroxyalkanoate synthase and impact on polyhydroxyalkanoate biosynthesis.

Aeromonas caviae polyhydroxyalkanoate synthase (PhaC(Ac)) is an important biocatalyst for the synthesis of practically useful two-component polyhydroxyalkanoate copolymer, poly[(R)-3-hydroxybutyrate-co-(R)-3-hydroxyhexanoate] [P(3HB-co-3HHx)]. In a previous study, two PhaC(Ac) mutants that have a single amino acid substitution of either asparagine 149 by serine (N149S) or aspartate 171 by glycine (D171G) were isolated as higher active enzymes by means of evolutionary engineering. In this study, the synergistic effects of N149S and D171G double mutation (NSDG) in PhaC(Ac) on polyhydroxyalkanoate biosynthesis were investigated in recombinant Ralstonia eutropha. The PhaC(Ac) NSDG mutant showed enhanced incorporation of longer 3-hydroxyalkanoate (3HA) units into the polyhydroxyalkanoate copolymer from octanoate (3HA fraction: 18.5 mol%) and soybean oil (5.4 mol%) as a carbon source. Besides, the NSDG mutant synthesized P(3HB) homopolymer with a very high molecular weight (M(w)=368 x 10(4)) when fructose was used as a carbon source. Thus, a combination of the beneficial mutations synergistically altered enzymatic properties, leading to synthesis of a polyhydroxyalkanoate copolymer with enhanced 3HA fraction and increased molecular weight.

Branched poly(lactide) synthesized by enzymatic polymerization: effects of molecular branches and stereochemistry on enzymatic degradation and alkaline hydrolysis.

In this article the effects of the number of molecular branches (chain ends) and the stereochemistry of poly(lactide)s (PLAs) on the enzymatic degradation and alkaline hydrolysis are studied. Various linear and branched PLAs were synthesized using lipase PS (Pseudomonas fluorescens)-catalyzed ring-opening polymerization (ROP) of lactide monomers having different stereochemistries (L-lactide, D-lactide, and D,L-lactide). Five different alcohols were used as initiators for the ROP, and the monomer-to-initiator molar feed ratio was varied from 10 to 100 and 1000 for each branch in the polymer architecture. The properties of branched PLAs that would affect the enzymatic and alkaline degradations, i.e., the glass transition temperature, the melting temperature, the melting enthalpy, and the advancing contact angle, were determined. The PLA films were degraded using proteinase K or 1.0 M NaOH solution, and the weight loss and changes in the number average molecular weight (Mn) of the polymer were studied during 12 h of degradation. The results suggest that an increase in the number of molecular branches of branched PLAs enhances its enzymatic degradability and alkali hydrolyzability. Moreover, the change in Mn of the branched poly(L-lactide) (PLLA) by alkaline hydrolysis indicated that the decrease in Mn was in the first place dependent on the number of molecular branches and thereafter on the length of the molecular branch of branched PLA. The branched PLLA, poly(D-lactide) (PDLA), and poly(D,L-lactide) (PDLLA) differed in weight loss and change in Mn of the PLA segment during the enzymatic degradation. It is suggested that the branched PDLLA was degraded preferentially by proteinase K.

Variation in copolymer composition and molecular weight of polyhydroxyalkanoate generated by saturation mutagenesis of Aeromonas caviae PHA synthase.

Amino acid substitutions at two residues downstream from the active-site histidine of polyhydroxyalkanoate (PHA) synthases are effective for changing the composition and the molecular weight of PHA. In this study, saturation mutagenesis at the position Ala505 was applied to PHA synthase (PhaCAc) from Aeromonas caviae to investigate the effects on the composition and the molecular weight of PHA synthesized in Ralstonia eutropha. The copolymer composition and molecular weight of PHA were varied by association with amino acid substitutions. There was a strong relationship between copolymer composition and PHA synthase activity of the cells. This finding will serve as a rationale for producing tailor-made PHAs.

Adsorption and hydrolysis reactions of poly(hydroxybutyric acid) depolymerases secreted from Ralstonia pickettii T1 and Penicillium funiculosum onto poly[(R)-3-hydroxybutyric acid].

Reaction processes of poly[(R)-3-hydroxybutyric acid] (P(3HB)) with two types of poly(hydroxybutyric acid) (PHB) depolymerases secreted from Ralstonia pickettii T1 and Penicillium funiculosum were characterized by means of atomic force microscopy (AFM) and quartz crystal microbalance (QCM). The PHB depolymerase from R. pickettii T1 consists of catalytic, linker, and substrate-binding domains, whereas the one from P. funiculosum lacks a substrate-binding domain. We succeeded in observing the adsorption of single molecules of the PHB depolymerase from R. pickettii T1 onto P(3HB) single crystals and the degradation of the single crystals in a phosphate buffer solution at 37 degrees C by real-time AFM. On the contrary, the enzyme molecule from P. funiculosum was hardly observed at the surface of P(3HB) single crystals by real-time AFM, even though the enzymatic degradation of the single crystals was surely progressed. On the basis of the AFM observations in air of the P(3HB) single crystals after the enzymatic treatments, however, not only the PHB depolymerase from R. pickettii T1 but also that from P. funiculosum adsorbed onto the surface of P(3HB) crystals, and both concentrations of the enzymes on the surface were nearly identical. This means both enzymes were adsorbed onto the surface of P(3HB) single crystals. Moreover, QCM measurements clarified quantitatively the differences in detachment behavior between two types of PHB depolymerases, namely the enzyme from R. pickettii T1 was hardly detached but the enzyme from P. funiculosum was released easily from the surface of P(3HB) crystals under an aqueous condition.

A human antibody fragment with high affinity for biodegradable polymer film.

Antibodies with high affinity for the surface of a solid material would be advantageous in biomaterial science as a protein device. A human antibody fragment that binds to poly(hydroxybutyrate) (PHB), a biodegradable polymer matter, was generated by a phage display system. Clone PH7-3d3 was isolated after several rounds of selection and prepared as a fragment of immunoglobulin variable regions (Fv). The quartz crystal microbalance technique showed that PH7-3d3 Fv completely inhibited PHB enzymatic degradation by competing with PHB depolymerase. Kinetic analysis based on surface plasmon resonance demonstrated that PH7-3d3 Fv bound to the PHB film with an equilibrium dissociation constant of 14 nM. The three-dimensional structure of PH7-3d3 Fv was resolved to 1.7 A, revealing that the complementarity determining regions (CDRs) in the Fv fragment form a relatively flat surface on which uncharged polar and aromatic amino acids are distributed in clusters. The structure of PH7-3d3 Fv was similar to that of PHB depolymerase in the orientation of aromatic residues in the binding sites. Alanine scanning mutagenesis demonstrated that these aromatic residues, especially tryptophan residues in CDRs, were critical in the interaction between PH7-3d3 Fv and PHB. Our results suggest the possible selection of an antibody fragment that binds a material surface in a manner similar to protein-ligand interaction.