Terminal alkyne formation by a pyridoxal phosphate-dependent enzyme

Area of Science:

• Biochemistry
• Enzymology
• Organic Chemistry

Background:

• Terminal alkynes are crucial building blocks in bio-orthogonal chemistry, particularly the Cu(I)-catalyzed azide-alkyne cycloaddition.
• The biosynthesis of L-β-ethynylserine involves the enzyme BesB, a pyridoxal phosphate-dependent enzyme, forming L-propargylglycine from a vinyl halide.
• Understanding the mechanism of such enzymatic reactions is key to expanding biocatalysis.

Purpose of the Study:

• To elucidate the catalytic mechanism of BesB in forming terminal alkynes.
• To present the high-resolution crystal structure of BesB.
• To explore the potential for developing novel biocatalysts for allene and alkyne formation.

Main Methods:

• X-ray crystallography to determine the 1.3-Å resolution structure of BesB.
• Detailed mechanistic studies involving substrate exchange reactions.
• Computational studies to support the proposed reaction pathway.

Main Results:

• The crystal structure of BesB was determined at 1.3-Å resolution.
• BesB was shown to reversibly catalyze halogen exchange in 4-halo-allyl-L-glycines.
• An allene intermediate was observed and implicated in the formation of the terminal alkyne via isomerization.

Conclusions:

• The mechanism involves deprotonation-driven halogen loss to form an allene intermediate, followed by isomerization to the terminal alkyne.
• This work expands the known catalytic capabilities of pyridoxal phosphate-dependent enzymes.
• The findings pave the way for designing metal-free biocatalysts for allene and alkyne synthesis.