Ligand binding and slow structural changes in chlorocruorin from Spirographis spallanzanii.
Chlorocruorin is a cooperative respiratory pigment found in the blood of polychaete worms; its prosthetic group is a derivative of the iron protoporphyrin IX, in which the vinyl group at position 2 is substituted by a formyl group. The quaternary structure of chlorocruorins is complex: myoglobin-like subunits are grouped in tetramers and tetramers in dodecamers; 12 dodecamers are assembled in the 3500-kDa particle. Chlorocruorin from Spirographis spallanzanii displays the following overall functional properties: (i) the oxygen affinity is lower than in human hemoglobin, while that of CO is similar if not higher; (ii) the rates of combination with oxygen and carbon monoxide are low; and (iii) the off rate of oxygen is comparable to that of human hemoglobin, while the off rate of CO is 10 times smaller. When CO is partially photolyzed with a long and powerful light flash (70 microseconds), rebinding is biphasic as in mammalian hemoglobins; however, the slowest rate is faster than that observed by stopped flow, suggesting that the unliganded protein decays from the liganded high affinity state (R) to an intermediate state before reaching the low affinity (T) state. Oxygen binding was followed by stopped-flow and flash photolysis. While partial photolysis yields a fast, second-order time course, stopped-flow experiments yield slow, biphasic, and non-second-order time courses. This pattern of reactivity was attributed to a slow conformational transition(s) which is (are) rare limiting with oxygen, but not with CO.(ABSTRACT TRUNCATED AT 250 WORDS)
Bellelli
A
A
Department of Biochemical Sciences Alessandro Rossi Fanelli, University of Rome, La Sapienza, Italy.
Lendaro
E
E
Ippoliti
R
R
Regan
R
R
Gibson
Q H
QH
Brunori
M
M
D000818:Animals; D001665:Binding Sites; D002248:Carbon Monoxide / Q000378:metabolism; D006420:Hemeproteins / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008024:Ligands; D010100:Oxygen / Q000378:metabolism*; D010782:Photolysis; D011077:Polychaeta; D011487:Protein Conformation
D006420:Hemeproteins; D008024:Ligands; C010884:chlorocruorin; D002248:Carbon Monoxide; D010100:Oxygen