pH Dependence and Cofactor Requirements of Photochemical Reactions in Maize Chloroplasts

Bishop, David G.; Andersen, Kirsten S.; Smillie, Robert M.

doi:10.1104/pp.50.6.774

Cited by 12 publications

(1 citation statement)

References 15 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In these metabolic processes, a series of soluble proteins/enzymes and membrane protein complexes play pivotal roles in electron transfer by communicating with target and binding molecules in response to changes in environmental conditions such as variations in pH, temperature, and the redox state [6][7][8].…”

Section: Introductionmentioning

confidence: 99%

Physicochemical nature of interfaces controlling ferredoxin NADP+ reductase activity through its interprotein interactions with ferredoxin

Kinoshita

Kim

Kume³

et al. 2015

Biochimica et Biophysica Acta (BBA) - Bioenergetics

View full text Add to dashboard Cite

Although acidic residues of ferredoxin (Fd) are known to be essential for activities of various Fd-dependent enzymes, including ferredoxin NADP(+) reductase (FNR) and sulfite reductase (SiR), through electrostatic interactions with basic residues of partner enzymes, non-electrostatic contributions such as hydrophobic forces remain largely unknown. We herein demonstrated that intermolecular hydrophobic and charge-charge interactions between Fd and enzymes were both critical for enzymatic activity. Systematic site-directed mutagenesis, which altered physicochemical properties of residues on the interfaces of Fd for FNR /SiR, revealed various changes in activities of both enzymes. The replacement of serine 43 of Fd to a hydrophobic residue (S43W) and charged residue (S43D) increased and decreased FNR activity, respectively, while S43W showed significantly lower SiR activity without affecting SiR activity by S43D, suggesting that hydrophobic and electrostatic interprotein forces affected FNR activity. Enzyme kinetics revealed that changes in FNR activity by mutating Fd correlated with Km, but not with kcat or activation energy, indicating that interprotein interactions determined FNR activity. Calorimetry-based binding thermodynamics between Fd and FNR showed different binding modes of FNR to wild-type, S43W, or S43D, which were controlled by enthalpy and entropy, as shown by the driving force plot. Residue-based NMR spectroscopy of (15)N FNR with Fds also revealed distinct binding modes of each complex based on different directions of NMR peak shifts with similar overall chemical shift differences. We proposed that subtle adjustments in both hydrophobic and electrostatic forces were critical for enzymatic activity, and these results may be applicable to protein-based electron transfer systems.

show abstract

Section: Introductionmentioning

confidence: 99%