From Residues to Added-Value Bacterial Biopolymers as Nanomaterials for Biomedical Applications

Abstract

Bacterial biopolymers are naturally occurring materials comprising a wide range of molecules with diverse chemical structures that can be produced from renewable sources following the principles of the circular economy. Over the last decades, they have gained substantial interest in the biomedical field as drug nanocarriers, implantable material coatings, and tissue-regeneration scaffolds or membranes due to their inherent biocompatibility, biodegradability into nonhazardous disintegration products, and their mechanical properties, which are similar to those of human tissues. The present review focuses upon three technologically advanced bacterial biopolymers, namely, bacterial cellulose (BC), polyhydroxyalkanoates (PHA), and γ-polyglutamic acid (PGA), as models of different carbon-backbone structures (polysaccharides, polyesters, and polyamides) produced by bacteria that are suitable for biomedical applications in nanoscale systems. This selection models evidence of the wide versatility of microorganisms to generate biopolymers by diverse metabolic strategies. We highlight the suitability for applied sustainable bioprocesses for the production of BC, PHA, and PGA based on renewable carbon sources and the singularity of each process driven by bacterial machinery. The inherent properties of each polymer can be fine-tuned by means of chemical and biotechnological approaches, such as metabolic engineering and peptide functionalization, to further expand their structural diversity and their applicability as nanomaterials in biomedicine.

Keywords: bacterial cellulose; bacterial polymers; biomedical applications; biopolymer functionalization; polyhydroxyalkanoates; upcycled polymers; γ-polyglutamic acid.

Figure 1

Chemical, microscopic, and macroscopic structure of BC, PHA, and PGA produced by model bacteria Komagataeibacter medellinensis, Pseudomonas putida, and Bacillus subtilis. Upper panels represent the chemical polymer structure, middle panels show electron microscopy images of the microorganisms producing the polymer, and lower panels show the macroscopic appearance of the purified polymer of BC (A), PHA (B), or PGA (C). SEM images of K. medellinensis and B. subtilis. Reprinted with permission from [14,15]; Copyright Microbiology Society, 2013, 2006.

Figure 2

Metabolic network of BC in Komagataeibacter xilynus E25. Sugars are metabolized through the pentoses phosphate pathway (PPP) to Glc-6-P, while glycolysis is not a relevant pathway in Komagataeibacter species due to the lack of phosphofructokinase. Glucose is partially oxidized in the periplasm to obtain reductor power. In the case of K. xylinus E25, the oxidation product is gluconic acid, although the final product is species-dependent. Ethanol is dehydrogenized to acetate by ADH1, an ADH2 inner-membrane-bound enzyme, and directed to Glc-6-P by the tricarboxylic acid cycle (TCA) and gluconeogenesis (GNG) pathways. Glc-6-P is isomerized to Glc-1-P by phosphoglucomutase (PGM) and is subsequently transformed to UDP-Glc by UTP–Glc-1-P uridylyltransferase (UGPT). Upon activation by c-di-GMP of BcsA, UDP-Glc units are polymerized into nascent glucan chains coupled with its translocation to the periplasm by means of cellulose synthase subunits BcsA (A) and BcsB (B). BcsC (C) is then involved in the arrangement of the nascent chains, and BcsD (D) forms the pore to export the nanofibrils. The main metabolic pathways, TCA, GNG, and PPP, are indicated in green. The pathway leading to BC synthesis is indicated in blue. Key enzymes, phosphoglucoisomerase (PGI), phosphoenol pyruvate carboxykinase (PEPCK), PGM, and UGPT are indicated. OAA: oxalacetate; PEP: phosphoenol pyruvate; 3PGA: 3-phosphoglycerate; GA3P: glyceraldehyde-3-P.

Figure 3

Metabolic network of PHA metabolism in model bacteria C. necator H16 (scl-PHA) and P. putida KT2440 (mcl-PHA). Fatty acids are metabolized via the β-oxidation cycle into acetyl-CoA, while nonfatty acid substrates are metabolized via the Entner-Doudoroff (ED) pathway, TCA and PPP, into acetyl-CoA. In PHB metabolism, two acetyl-CoA molecules condensate into acetoacetyl-CoA by PhaB and are then converted into R-3-hydroxybutyryl-CoA (PHB monomer) by PhaA. In PHA metabolism, fatty acids are metabolized in the β-oxidation cycle, where the intermediates 3-ketoacyl-CoA and trans-2-enoyl-CoA can be directly converted into PHA monomers (R-3-HA-CoA) by FabG and PhaJ, respectively. Alternatively, through de novo fatty acid synthesis, acetyl-CoA can be converted from R-3-hydroxyl-ACP to R-3-HA-CoA by two enzymatic steps catalyzed by PhaG and AlkK. PHA and PHB are synthetized in a continuous cycle that drives carbon and energy flux, in which the monomers are polymerized by PhaC, depolymerized into the respective R-3-hydroxycarboxylic acids by PhaZ, and reconverted into the activated monomer R-3-HA-CoA by Acs1. The carbon central metabolic pathways are indicated in green, the pathway leading to PHB synthesis is in purple, and the one leading to PHA synthesis is shown in blue. Key enzymes are indicated.

Figure 4

Metabolic network of PGA metabolism in model bacteria B. subtilis 168. Exogenous glu can serve as a direct precursor of PGA synthesis. Alternatively, sugars are metabolized via the Embden Meyerhof (EM) pathway, PPP and TCA. αKG from TCA is then converted to L-glu (the PGA monomer) by two different enzymes—glutamine oxoglutarate aminotransferase (GOGAT), which transfers the amino group from a glutamine molecule, and Glu dehydrogenase (GluDH), which incorporates the amino group from an ammonium molecule. In the species producing L/D-PGA (such as B. subtilis), a racemization reaction to produce the D-isomer takes place. This can be produced by means of two different enzymatic reactions: the Glu racemase (RacE) directly interconverting the isomers and a 3-enzymatic-step reaction by L-glu-pyruvate aminotrasnferase (L-Glu-AT), alanine racemase (Alr), and D-glu-pyruvate aminotransferase (D-Glu-AT). D/L glu monomers are then polymerized in the active site formed by membrane-bound PgsB (B) and PgsC (C), and the elongated chain is then removed from the active site by PgsA (A). The role of PgsE (E) is still under debate, while PgdS (S) is a secreted peptidase that releases the PGA to the medium. The main metabolic pathways, TCA, GNG, and PPP, are indicated in green. The pathway leading to PGA synthesis is indicated in blue. Key enzymes, glucokinase (GK), GOGAT, GluDH, glutamine synthetase (GS), L-Glu-AT, Alr, D-Glu-AT, and RacE are indicated.

Figure 5

Flowchart of industrial and municipal upcycling of residues into bacterial polymers. Box colors indicate the highest percentage of carbon source in composition: green (saccharides), yellow (lipids, fatty acids), light grey (aminoacids), and dark grey (recalcitrant compounds: syngas, CO₂, aromatics, and BTEXS. BTEXS includebenzene, ethylbenzen, toluene and xylene. Processes or treatments transforming the raw residues into bacteria assimilable substrates are indicated.

Figure 6

Flowchart of the different approaches used in the literature to modify bacterial polymers.