In Time for Michael Behe's Book Anniversary, Here's a Real Mousetrap in the Cell
Evolution News & Views
Later this year will mark the twentieth anniversary of Michael Behe's Darwin's Black Box, a book that gave profound impetus to the burgeoning intelligent design movement. With that in mind, we want to highlight examples of discoveries that vindicate Behe's arguments in the book. Today, here is one buried in a scientific paper that might have been missed. It's especially delightful because it brings to life an analogy Behe made famous: the mousetrap as an example of irreducible complexity.Dr. Behe defined irreducible complexity as "a single system composed of several well-matched, interacting parts that contribute to the basic function, wherein the removal of any one of the parts causes the system to effectively cease functioning" (p. 39). A few pages later, he explained how the "humble mousetrap" meets the requirements of an irreducibly complex system: it has a function (catching mice), and all five parts (base, spring, hammer, catch and holding bar) are necessary for that function.Behe had said that "An irreducibly complex biological system, if there is such a thing, would be a powerful challenge to Darwinian evolution." In subsequent chapters, he provided many examples in nature, from molecular machines in the cell like the cilium and flagellum, to whole-body systems like the blood clotting cascade.
Now a new example: a molecular machine that works like a mousetrap. How cool is that? No kidding, here's how the authors of a paper in the Proceedings of the National Academy of Sciences describe serpin antithrombin III (for short, ATIII):
Here the cellular folding pathway of the serpin antithrombin III (ATIII), which inhibits proteases involved in the coagulation cascade, was determined. ATIII uses a large conformational movement in a mousetrap-like mechanism to bind and distort its target protease, resulting in protease inhibition. This work establishes that folding to an active, cocked state requires early stabilization of the C-terminal region, which is the last sequence translated, explaining how the serpin or mousetrap is set. [Emphasis added.]
Oh, this is good. Here we find a mousetrap, a molecular machine, and the blood clotting cascade brought together in a single irreducibly complex protein. And proteins, we all know, are coded by complex specified information -- another hallmark of intelligent design -- in the genome.
Be glad you have ATIII in your bloodstream. It's an anticoagulant, helping prevent thrombosis and pulmonary embolism. It plays a very important role in regulating normal blood coagulation. As a serpin (serine protease inhibitor), its job is to prevent runaway clotting by deactivating a certain protease called thrombin. Behe actually talks about it in his book (p. 85) in the chapter about the blood clotting cascade:
Once clotting has begun, what stops it from continuing until all the blood in the animal as solidified? Clotting is confined to the site of injury in several ways. (Please refer to Figure 4-3.) First, a plasma protein called antithrombin binds to the active (but not the inactive) forms of most clotting proteins and inactivates them. Antithrombin is itself relatively inactive, however, unless it binds to a substance called heparin. Heparin occurs inside cells and undamaged blood vessels.
From there, the complexity of the blood clotting system rises dramatically. Antithrombin (ATIII being the principal form) is thus a key player in this example of what Behe demonstrated to be an irreducibly complex (IC) biological system. In 1996, ATIII's structure was little known. Now, seven molecular biologists are telling us it works like a mousetrap!
But can you be sure ATIII is itself irreducibly complex? First, note that the seven authors of the PNAS paper, all from the University of Massachusetts, never explain how this protein might have evolved. Quite the contrary; their only mention of "evolution" deals with how the protein folds, not with Darwinian evolution. There's no mention of selection, phylogeny, or ancestors. Instead, they seem fascinated by the precise way this machine must be assembled and "cocked" for action. Watch for "mousetrap" again:
Irreversible switching from one conformation to another allows proteins to perform mechanical work without external energy sources such as ATP. Large conformational movements, up to ∼100 Å, can be triggered by proteolysis or changes in environmental conditions, such as pH, initiating processes including membrane fusion for viral infection, protease activation, or inhibition. To facilitate these processes, proteins must fold to kinetically trapped metastable states with relatively high free energy. How proteins fold to these states and avoid more thermodynamically stable conformations is poorly understood. The serpin family of serine protease inhibitors exemplifies this type of metastable protein, and its mechanism of folding presents a conundrum. The native, active serpin fold positions the target protease-binding site on a loosely structured, accessible stretch of sequence termed the reactive center loop (RCL). Once the protease forms a covalent acyl intermediate in the scissile bond in the RCL and cleaves the bond, the serpin undergoes a major conformational change like the springing of a mousetrap, and the protease is carried ∼70 Å to the opposite side of the serpin, thereby inactivating the protease by mechanical deformation (Fig. S1). Strikingly, the conformational landscape of serpins has an alternate fold that is more stable than the functionally required "cocked mousetrap" fold. In this alternative fold, called the latent state, the intact RCL is inserted as an additional strand into the central β-sheet, resulting in a more stable but inactive state.
Are we having fun yet? Here is a molecular machine that must be cocked like a mousetrap, storing energy for its function. In a very real sense, the "mice" that ATIII needs to catch are proteases. Just as a sprung mousetrap is in a more stable state but can't catch mice, ATIII has a more stable thermodynamic state but can't catch proteases. To latch onto and deactivate proteases, ATIII must first be cocked by a precisely operated folding sequence and association with heparin. Only then, when it finds its prey, it "undergoes a major conformational change like the springing of a mousetrap" to deactivate it.
And thus you are kept healthy, neither bleeding to death nor clotting to death. The irreducible complexity of ATIII is also demonstrated by showing what happens when it fails. They found that out by looking at a variety of mutant forms. Keep in mind that in an IC system each part is necessary for function:
Encoding this gymnastic ability in the folding landscape of serpins comes at a risk: Many mutations in serpins cause misfolding and are associated with diseases called serpinopathies.
The serpin antithrombin III (ATIII) plays an essential role in blood clotting by regulating the activity of thrombin and other serine proteases in the coagulation cascade. Numerous misfolding mutations of ATIII are linked to thrombosis. The cellular folding process of ATIII, including its traversal of the secretory pathway, facilitates its folding to the functional metastable high free energy state. The differing outcomes of unassisted refolding of purified ATIII and its cellular folding underline the profound difference between protein folding reactions in isolation and in cells and invite further exploration of the key players and steps that make cellular folding so successful. The fact that ATIII and other serpins must adopt metastable states to function and that their misfolding is implicated in several pathologies further raises the importance of understanding their cellular folding pathway.
The details of the folding pathway need not concern us here, except to note that they are precise and ordered. The easiest thing for this protein to do after it emerges from the ribosome would be to fold into a stable clump that does nothing. That's what happens in test tubes. The reason functional ATIII folds "rapidly and efficiently" in cells, the scientists found, is because it proceeds in a sequence of steps, aided by chaperones. They suspected a conspiracy at work:
We hypothesized that the ER carbohydrate-binding chaperones calnexin and calreticulin may be coconspirators in high-fidelity cellular folding of ATIII. We tested this hypothesis by treatment of ATIII producing cells with the glucosidase inhibitor castanospermine (CST), which prevents the formation of monoglucosylated glycoproteins and thereby inhibits lectin chaperone binding. In the presence of CST, the level of ATIII secreted was reduced by a factor of three (Fig. 3A, lanes 28-36). The activity of the secreted triglucosylated protein was also modestly lower than untreated ATIII (Fig. 3B). Together, these results demonstrate the importance of glucosidase trimming and subsequent lectin chaperone binding for the efficient maturation of functional ATIII.
That gives a modest sense of the overarching lesson here: multiple factors are working together to make ATIII work. This machine, in turn, requires genetic instructions that exhibit specified complexity, or else function is lost. And ATIII is one of multiple factors working together to make blood clotting functional. As biologist Jonathan Wells remarked in Unlocking the Mystery of Life, "What we have here is irreducible complexity all the way down."
Evolution News & Views
Later this year will mark the twentieth anniversary of Michael Behe's Darwin's Black Box, a book that gave profound impetus to the burgeoning intelligent design movement. With that in mind, we want to highlight examples of discoveries that vindicate Behe's arguments in the book. Today, here is one buried in a scientific paper that might have been missed. It's especially delightful because it brings to life an analogy Behe made famous: the mousetrap as an example of irreducible complexity.Dr. Behe defined irreducible complexity as "a single system composed of several well-matched, interacting parts that contribute to the basic function, wherein the removal of any one of the parts causes the system to effectively cease functioning" (p. 39). A few pages later, he explained how the "humble mousetrap" meets the requirements of an irreducibly complex system: it has a function (catching mice), and all five parts (base, spring, hammer, catch and holding bar) are necessary for that function.Behe had said that "An irreducibly complex biological system, if there is such a thing, would be a powerful challenge to Darwinian evolution." In subsequent chapters, he provided many examples in nature, from molecular machines in the cell like the cilium and flagellum, to whole-body systems like the blood clotting cascade.
Now a new example: a molecular machine that works like a mousetrap. How cool is that? No kidding, here's how the authors of a paper in the Proceedings of the National Academy of Sciences describe serpin antithrombin III (for short, ATIII):
Here the cellular folding pathway of the serpin antithrombin III (ATIII), which inhibits proteases involved in the coagulation cascade, was determined. ATIII uses a large conformational movement in a mousetrap-like mechanism to bind and distort its target protease, resulting in protease inhibition. This work establishes that folding to an active, cocked state requires early stabilization of the C-terminal region, which is the last sequence translated, explaining how the serpin or mousetrap is set. [Emphasis added.]
Oh, this is good. Here we find a mousetrap, a molecular machine, and the blood clotting cascade brought together in a single irreducibly complex protein. And proteins, we all know, are coded by complex specified information -- another hallmark of intelligent design -- in the genome.
Be glad you have ATIII in your bloodstream. It's an anticoagulant, helping prevent thrombosis and pulmonary embolism. It plays a very important role in regulating normal blood coagulation. As a serpin (serine protease inhibitor), its job is to prevent runaway clotting by deactivating a certain protease called thrombin. Behe actually talks about it in his book (p. 85) in the chapter about the blood clotting cascade:
Once clotting has begun, what stops it from continuing until all the blood in the animal as solidified? Clotting is confined to the site of injury in several ways. (Please refer to Figure 4-3.) First, a plasma protein called antithrombin binds to the active (but not the inactive) forms of most clotting proteins and inactivates them. Antithrombin is itself relatively inactive, however, unless it binds to a substance called heparin. Heparin occurs inside cells and undamaged blood vessels.
From there, the complexity of the blood clotting system rises dramatically. Antithrombin (ATIII being the principal form) is thus a key player in this example of what Behe demonstrated to be an irreducibly complex (IC) biological system. In 1996, ATIII's structure was little known. Now, seven molecular biologists are telling us it works like a mousetrap!
But can you be sure ATIII is itself irreducibly complex? First, note that the seven authors of the PNAS paper, all from the University of Massachusetts, never explain how this protein might have evolved. Quite the contrary; their only mention of "evolution" deals with how the protein folds, not with Darwinian evolution. There's no mention of selection, phylogeny, or ancestors. Instead, they seem fascinated by the precise way this machine must be assembled and "cocked" for action. Watch for "mousetrap" again:
Irreversible switching from one conformation to another allows proteins to perform mechanical work without external energy sources such as ATP. Large conformational movements, up to ∼100 Å, can be triggered by proteolysis or changes in environmental conditions, such as pH, initiating processes including membrane fusion for viral infection, protease activation, or inhibition. To facilitate these processes, proteins must fold to kinetically trapped metastable states with relatively high free energy. How proteins fold to these states and avoid more thermodynamically stable conformations is poorly understood. The serpin family of serine protease inhibitors exemplifies this type of metastable protein, and its mechanism of folding presents a conundrum. The native, active serpin fold positions the target protease-binding site on a loosely structured, accessible stretch of sequence termed the reactive center loop (RCL). Once the protease forms a covalent acyl intermediate in the scissile bond in the RCL and cleaves the bond, the serpin undergoes a major conformational change like the springing of a mousetrap, and the protease is carried ∼70 Å to the opposite side of the serpin, thereby inactivating the protease by mechanical deformation (Fig. S1). Strikingly, the conformational landscape of serpins has an alternate fold that is more stable than the functionally required "cocked mousetrap" fold. In this alternative fold, called the latent state, the intact RCL is inserted as an additional strand into the central β-sheet, resulting in a more stable but inactive state.
Are we having fun yet? Here is a molecular machine that must be cocked like a mousetrap, storing energy for its function. In a very real sense, the "mice" that ATIII needs to catch are proteases. Just as a sprung mousetrap is in a more stable state but can't catch mice, ATIII has a more stable thermodynamic state but can't catch proteases. To latch onto and deactivate proteases, ATIII must first be cocked by a precisely operated folding sequence and association with heparin. Only then, when it finds its prey, it "undergoes a major conformational change like the springing of a mousetrap" to deactivate it.
And thus you are kept healthy, neither bleeding to death nor clotting to death. The irreducible complexity of ATIII is also demonstrated by showing what happens when it fails. They found that out by looking at a variety of mutant forms. Keep in mind that in an IC system each part is necessary for function:
Encoding this gymnastic ability in the folding landscape of serpins comes at a risk: Many mutations in serpins cause misfolding and are associated with diseases called serpinopathies.
The serpin antithrombin III (ATIII) plays an essential role in blood clotting by regulating the activity of thrombin and other serine proteases in the coagulation cascade. Numerous misfolding mutations of ATIII are linked to thrombosis. The cellular folding process of ATIII, including its traversal of the secretory pathway, facilitates its folding to the functional metastable high free energy state. The differing outcomes of unassisted refolding of purified ATIII and its cellular folding underline the profound difference between protein folding reactions in isolation and in cells and invite further exploration of the key players and steps that make cellular folding so successful. The fact that ATIII and other serpins must adopt metastable states to function and that their misfolding is implicated in several pathologies further raises the importance of understanding their cellular folding pathway.
The details of the folding pathway need not concern us here, except to note that they are precise and ordered. The easiest thing for this protein to do after it emerges from the ribosome would be to fold into a stable clump that does nothing. That's what happens in test tubes. The reason functional ATIII folds "rapidly and efficiently" in cells, the scientists found, is because it proceeds in a sequence of steps, aided by chaperones. They suspected a conspiracy at work:
We hypothesized that the ER carbohydrate-binding chaperones calnexin and calreticulin may be coconspirators in high-fidelity cellular folding of ATIII. We tested this hypothesis by treatment of ATIII producing cells with the glucosidase inhibitor castanospermine (CST), which prevents the formation of monoglucosylated glycoproteins and thereby inhibits lectin chaperone binding. In the presence of CST, the level of ATIII secreted was reduced by a factor of three (Fig. 3A, lanes 28-36). The activity of the secreted triglucosylated protein was also modestly lower than untreated ATIII (Fig. 3B). Together, these results demonstrate the importance of glucosidase trimming and subsequent lectin chaperone binding for the efficient maturation of functional ATIII.
That gives a modest sense of the overarching lesson here: multiple factors are working together to make ATIII work. This machine, in turn, requires genetic instructions that exhibit specified complexity, or else function is lost. And ATIII is one of multiple factors working together to make blood clotting functional. As biologist Jonathan Wells remarked in Unlocking the Mystery of Life, "What we have here is irreducible complexity all the way down."
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