Analysis of structure-function relationships in citrate lyase isolated from Rhodopseudomonas gelatinosa as revealed by cross-linking and immunoelectron microscopy.
Citrate lyase (EC 4.1.3.6) isolated from Rhodopseudomonas gelatinosa was analyzed using cross-linking experiments and immunoelectron microscopy. Different cross-linking reagents and antibodies directed against citrate lyase and specifically against all three subunit types (L, M and S) were applied. A structure-function model is proposed for citrate lyase from R. gelatinosa: the enzyme occurs in two configurations, 'rings' and 'stars'. The ring contains two identical layers each consisting of three subunits L, with one subunit S as a polar cap sitting on each L, and three subunits M in alternating sequence (18 subunits altogether). In the star, the same 18 subunits are arranged in a different way. Whereas the subunits L are located at the periphery, the subunits M are concentrated in the center of the particle. The subunits S are positioned relative to L as in the ring; however, their location relative to the subunits M is changed. By transition from ring to star, areas on S are brought into contact with areas on M by rotation of structural units, consisting of one L, one M and one S subunit per layer, against each other, with S of one structural unit close to M of the neighbouring structural unit. This transition is assumed to work also in reversed direction. The observation of rings and stars as two distinct molecular forms is proposed to reflect the two states of citrate lyase, the ring being the form where substrate is bound by acyl exchange, and the star being the form where the substrate is consumed by cleavage, i.e. the catalyzed reaction is completed.
D003432:Cross-Linking Reagents; D007120:Immunochemistry; D008854:Microscopy, Electron; D009097:Multienzyme Complexes / Q000302:isolation & purification*; D007652:Oxo-Acid-Lyases / Q000302:isolation & purification*; D012241:Rhodopseudomonas / Q000201:enzymology*; D013329:Structure-Activity Relationship
D003432:Cross-Linking Reagents; D009097:Multienzyme Complexes; D007652:Oxo-Acid-Lyases; C019909:citrate (pro-3S)-lyase