2025 Summer Research Symposium • Julia Martinez • July 9, 2025

Julia Martinez

Class of 2025

Majors: Forensic Science (Criminology)

Mentor: Audrey Lamb, PhD, University of Texas at San Antonio

Detection of Catalytic Intermediates in the RuBisCO Mechanism using 13C

and 31P NMR

Ribulose 1,5-bisphosphate carboxylase/oxygenase, or RuBisCO, is an enzyme which plays an

essential role in carbon fixation and plant metabolism. While RuBisCO is considered the most

abundant protein on the planet, its catalytic mechanism is still not completely understood.

RuBisCO is notoriously slow and exhibits low specificity for CO2 over O2, leading to

energetically wasteful photorespiration. Efforts to improve RuBisCO’s catalytic efficiency have

been hindered due to an incomplete picture of key transient intermediates involved in the

reaction. Acid quenched single turnover reaction experiments in combination with 13C and 31P

Nuclear Magnetic Spectroscopy (NMR) offers a promising approach for the characterization of

RuBisCO’s reaction pathway at a molecular level, including the detection of short-lived

intermediates. In this study, we present an initial framework for applying NMR techniques to

probe the RuBisCO catalytic pathway. We assess the feasability of detecting catalytic

intermediates under varying experimental conditions, including cofactor availability and reaction

time. Our preliminary data reveal both the promise and the technical challenges of this approach,

including issues related to sensitivity, intermediate lifetimes, and sample conditions. While the

direct observation of individual intermediates remains a target for future work, these findings

demonstrate the potential of NMR as a tool for investigating RuBisCO’s catalytic mechanism

and provide a technical foundation for future research aimed at the characterization of reaction

intermediates.

Keywords: Rubisco, enzyme, nuclear magnetic resonance, carbon fixation, intermediates