Substrate-binding model of the chlorophyll biosynthetic magnesium chelatase BchH subunit.
Photosynthetic organisms require chlorophyll and bacteriochlorophyll to harness light energy and to transform water and carbon dioxide into carbohydrates and oxygen. The biosynthesis of these pigments is initiated by magnesium chelatase, an enzyme composed of BchI, BchD, and BchH proteins, which catalyzes the insertion of Mg(2+) into protoporphyrin IX (Proto) to produce Mg-protoporphyrin IX. BchI and BchD form an ATP-dependent AAA(+) complex that transiently interacts with the Proto-binding BchH subunit, at which point Mg(2+) is chelated. In this study, controlled proteolysis, electron microscopy of negatively stained specimens, and single-particle three-dimensional reconstruction have been used to probe the structure and substrate-binding mechanism of the BchH subunit to a resolution of 25A(.) The apo structure contains three major lobe-shaped domains connected at a single point with additional densities at the tip of two lobes termed the "thumb" and "finger." With the independent reconstruction of a substrate-bound BchH complex (BchH.Proto), we observed a distinct conformational change in the thumb and finger subdomains. Prolonged proteolysis of native apo-BchH produced a stable C-terminal fragment of 45 kDa, and Proto was shown to protect the full-length polypeptide from degradation. Fitting of a truncated BchH polypeptide reconstruction identified the N- and C-terminal domains. Our results show that the N- and C-terminal domains play crucial roles in the substrate-binding mechanism.
Sirijovski
Nickolche
N
Department of Biochemistry, Center for Molecular Protein Science, Lund University, SE-22100 Lund, Sweden.
Lundqvist
Joakim
J
Rosenbäck
Matilda
M
Elmlund
Hans
H
Al-Karadaghi
Salam
S
Willows
Robert D
RD
Hansson
Mats
M
D001426:Bacterial Proteins / Q000737:chemistry*; D002734:Chlorophyll / Q000737:chemistry*; D008190:Lyases / Q000737:chemistry* / Q000378:metabolism; D008854:Microscopy, Electron; D008958:Models, Molecular; D008968:Molecular Conformation; D009532:Nickel / Q000737:chemistry; D010788:Photosynthesis; D011166:Porphyrins / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011524:Protoporphyrins / Q000737:chemistry; D016354:Rhodobacter capsulatus / Q000378:metabolism; D013379:Substrate Specificity
D001426:Bacterial Proteins; D011166:Porphyrins; D011524:Protoporphyrins; D002734:Chlorophyll; D009532:Nickel; C028025:protoporphyrin IX; D008190:Lyases; C086788:magnesium chelatase