User:Mitchell Long/Sandbox 1

Introduction

Luciferases are a class of enzymes that catalyze the oxidation of a long chain aliphatic aldehydes. This reaction results in the formation of a carboxylic acid and the emission of photons in the form of blue-green light. The luciferase found in 'Vibrio harveyi' is a heterodimer that is composed of a catalytic α subunit and a homologous but noncatalytic β subunit. The catalytic α subunit houses the FMN cofactor and is connected to the β subunit via a hairpin structure called the "mobile loop."

Mechanism

Luciferase found inV. Harveyi binds noncovalently to a reduced flavin mononucleotide cofactor, an aliphatic aldehyde and oxygen to yield oxidized flavin mononucleotide, water, and carboxylic acid. The reaction occurs in two steps forming a hydroxyflavin intermediate and ultimately results in the oxidation of the aldehyde and emission of photons in the form of blue green light.

FMNH₂+O₂+RCHO→FMN+RCOOH+H₂O+hv(490nm)

The catalytic α subunit houses the FMN cofactor and is connected to the β subunit via a hairpin structure called the "." The organic substrate for bacterial luciferase in vivo is myristic aldehyde, although many aliphatic aldehydes of various lengths can induce bioluminescence in vitro.

Structural Motifs

Structure homology-There is a great deal of sequence homology and structural coservation between the α and β subunits. When superimposed over eachother the barrels of the alpha and beta subunits with a deviation of 0.62Å for 42 equivalent α carbons. The region of the beta subunit that contains the 29 residue deletion with respect to the alpha subunit differs notably in arrangement. In the alpha subunit, the α7a helix is straight and extends toward the beta subunit. The region involved with dimerization, helices α and β and the hairpin loop structure are exceptionally similar in superposition.

Active Site and Alpha Subunit-the of bacterial luciferase is a large open cavity that is accessible to solvent via an opening located at the C-terminal ends of the ǰ strans of the TIM-barrel structure. During the first step of the oxidation reaction, FMNH₂ binds to the flavin binding pocket and the enzyme undergoes a conformational change that blocks water in the surrounding environment from accessing both the excited peroxydihydroflavin intermediate. Next O₂ and a long chain aldehyde bind to the FMNH₂ luciferase complex and a two step oxidatino reaction occurs. .

The β subunit-The beta subunit is characterized as a necessary but non-catalytic subunit that stabilizes the catalytic ǯ subunit that is responsible for the oxidation reaction. Both subunits share a great deal of homology

Mobile Loop- Phe2772-thr 288 (β/α)₈ Barrel- The tertiary structure of the α and β subunits is very similar. both subunits fold into a single-domain eight-stranded β/α barrel motif. the two subunits assemble around a parallel four-helix bundle centered on a pseudo 2-fold axis that relates the alpha and beta subunits. .