Assembly of the cysteine synthase complex and the regulatory role of protein-protein interactions

Sarangaliram Kumaran, Yi Hankuil, Hari B. Krishnan, Joseph M. Jez

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63 Scopus citations


Macromolecular assemblies play critical roles in regulating cellular functions. The cysteine synthase complex (CSC), which is formed by association of serine O-acetyltransferase (SAT) and O-acetylserine sulfhydrylase (OASS), acts as a sensor and modulator of thiol metabolism by responding to changes in nutrient conditions. Here we examine the oligomerization and energetics of formation of the soybean CSC. Biophysical examination of the CSC by size exclusion chromatography and sedimentation ultracentrifugation indicates that this assembly (complex Mr ∼ 330,000) consists of a single SAT trimer (trimer Mr ∼ 110,000) and three OASS dimers (dimer Mr ∼ 70,000). Analysis of the SAT-OASS interaction by isothermal titration calorimetry reveals negative cooperativity with three distinct binding events during CSC formation with Kd values of 0.3, 7.5, and 78 nM. The three binding events are also observed using surface plasmon resonance with comparable affinities. The stability of the CSC derives from rapid association and extremely slow dissociation of OASS with SAT and requires the C terminus of SAT for the interaction. Steady-state kinetic analysis shows that CSC formation enhances SAT activity and releases SAT from substrate inhibition and feedback inhibition by cysteine, the final product of the biosynthesis pathway. Cysteine inhibits SAT and the CSC with Ki values of 2 and 70μM, respectively. These results suggest a new model for the architecture of this regulatory complex and additional control mechanisms for biochemically controlling plant cysteine biosynthesis. Based on previous work and our results, we suggest that OASS acts as an enzyme chaperone of SAT in the CSC.

Original languageEnglish
Pages (from-to)10268-10275
Number of pages8
JournalJournal of Biological Chemistry
Issue number15
StatePublished - Apr 10 2009


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