Bioluminescence and secondary structure properties of aequorin mutants produced for site-specific conjugation and immobilization

J. C. Lewis, J. J. López-Moya, S. Daunert

Research output: Contribution to journalArticlepeer-review

23 Scopus citations


Aequorin is one of several photoproteins that emits visible light upon binding to calcium ions. It has been widely used as a Ca2+-indicator and as an alternative highly sensitive bioluminescent label in binding assays. The apoprotein of aequorin binds an imidazopyrazine compound (coelenterazine) and molecular oxygen to form a stable photoprotein complex. Upon addition of calcium, the photoprotein undergoes a conformational change leading to the oxidation of the chromophore with the release of CO2 and blue light. To gain more information of structure-function relationships within the photoprotein that will aid in the design of mutants suitable for site-specific conjugation and immobilization, polymerase chain reaction (PCR)-based site-directed mutagenesis was employed to produce five different aequorin mutants. The five mutants included a cysteine-free mutant and four other mutants with single cysteine residues at selected positions within the protein. The aequorin mutants exhibited different bioluminescence emission characteristics with two mutants showing a decrease in relative light production in comparison to the cysteine-free mutant. Additionally, circular dichroism (CD) spectra revealed that the single amino acid substitutions made for two of the aequorin mutants did alter their secondary structures.

Original languageEnglish (US)
Pages (from-to)65-70
Number of pages6
JournalBioconjugate Chemistry
Issue number1
StatePublished - Jan 2000
Externally publishedYes

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Pharmacology
  • Pharmaceutical Science
  • Organic Chemistry


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