Metal ion adsorption is dependent upon the substrate pH value. When the pH is acidic, removal rates are typically much lower, a trend resulting from the ionisation of functional groups present on the mycelium cell surface (electrically neutral atoms are converted to electrically charged atoms). Lower removal rates in acidic conditions could also be a consequence of solution protonation. In this latter event, the surge in protons increases competition for negative binding sites on the cell surface which reduces their availability for Ni(II) ions. As the pH of the solution or substrate increases, the competition for binding sites depreciates, enabling the observed escalation in Ni(II) removal rates. At the optimum pH, of 5-6, electrostatic interactions towards positively charged Ni(II) ions are generated by fungal strains. Beyond this optimal range, binding affinity weakens in line with the ion sink created by metal complexes and hydroxides.

Enzymatic antioxidants:

• Peroxidase; POD, 

• superoxide-dismutase; SOD, 

• catalase; CAT, 

• polyphenol oxidase; PPO

Fungal cell walls contain a rich arsenal of polysaccharides, lipids and proteins that accommodate the electrostatic interactions by which heavy metals are sequestered from contaminated substrates.  One such polysaccharide is chitin, whose polar functional groups - carboxyl, amino, hydroxyl - are responsible for the chemisorption of Ni(II). 

Fungi require trace amounts of metals for normal metabolic processes. At low concentrations, metal ions can produce stimulatory responses in fungal biomass. Sharma et al (2020) demonstrated this preference for substrates containing low concentrations of metals by the weight of fungal biomass. At low concentrations of Pb, Cd and Ni, fungal biomass grew to 0.67 g l−1, while in the absence of metals, they grew to a mere 0.24g 1-1. Beyond these low concentrations, metal ions tend to prove toxic for most fungal species with the exception of strains possessing directed metabolic adaptions that facilitate inhibition, transformation and tolerance of metal ions. 

Functional Groups: 

• Carbonyl

• Amino

• Hydroxyl

• Phosphate

• Nitro

• ATP-binding cassette (regulates the import and export of substances across plasma membranes) intracellular transfer of heavy metals and further subcellular compartmentation

• Detoxification either by transport into the vacuole or export from the cell through Mte1 of the MATE transporter family (Tricholoma vaccinum - Schlunk et al 2015)

• Ni transport through Ni/Co (nickel/cobalt) transporters of ABC2 importers family (previously reported in Schizosaccharomyces pombe by (Eitinger et al. 2000))

• Bpt1p, Ycf1p, Vmr1p and Nft1p multidrug resistance-associated proteins in transport heavy metals (Cd) and GSH conjugates from the cytosol into the vacuole in Saccharomyces cerevisiae (Sipos and Kuchler 2006)

• In S.cerevisiae and S.pombe, the transfer of phytochelatin-Cd and GSH-Pb complexes into the vacuole is mediated by Hmt1p and Ycf1p. 

• Cation Diffuse Facilitator and Zrt/Irt-like protein has been identified as a Zn transporter in fungi but could also play a role in the transport of other divalent metal ions such as Ni, even if to a lesser extent. If two chemically similar metals are present, one might be able to replace the other in a transport system (import or export pathways) (as is the case with copper/zinc in plants), resulting in either its accumulation or efflux. 

• HMS (heavy-metal stress) provokes ROS production. ROS left untreated will negatively react with lipids, proteins, etc resulting in enzyme inhibition or inactivation, and membrane damage. ROS triggers the production of thiol compounds (Metallothionein-MT and glutathione-GSH). Intracellular production of MT and GSH can promote metal ion chelation, thus enabling subcellular compartmentation. These thiol compounds have been similarly implicated in the binding and accumulation of metal ions into intracellular spaces and the vacuole, resulting in increased tolerance and lower cytoplasmic toxicity (Damodaran et al 2013) The stoichiometric values of ROS and thiol compounds are strong determinants of metal resistance. 

• H202 (ROS) increases the production and concentration of melanodialdehydes, the marker of membrane lipid peroxidation, in A.flvus Cr700 by 277% under Ni stress

• at 100mg/L of Ni, POD activity increased by 679.5%

• Changes in the activities of CAT, POD, and SOD are in direct correspondence with the changes seen in MDA and H202 under metal stress; which points to the importance of antioxidant enzymes in scavenging reactive oxygen species. Superoxide-dismutase converts 02- into O2; while h202 is converted to h2o by CAT. While enzymatic systems are in full operation, non-enzymatic allies are reinforcing antioxidant efforts. Proline is one such antioxidant that is produced under abiotic stresses including adverse environmental conditions and heavy metal exposure. Proline acts as a support system by stabilizing proteins and antioxidant enzymes, as well as balancing intracellular redox homeostasis. Phenolic compounds are another defense against oxidative stress. Phenolic compounds decrease ROS formation by inhibiting enzymes involved in their generation and upregulation.