Thursday, December 30, 2010

671)The Bacterial Flagellum – Truly An Engineering Marvel!; Quotes From Blogpost Four Hundred.

"Every particle of the Creation has a share of the Command of God, because every creature shares a part of the Command of God through which it has come to be there and by virtue of which it remains in being and the light of the Command ofGod shines in it. Understand this!"(Abu Yakub Al Sijistani, 10th century Fatimid Ismaili cosmologist, d971, Kashf al-Mahjub("Unveiling of the Hidden"))

Kathalika yubayyinu Allahu lakum ayatihi la'allakum ta-'aqiloona: "Allah thus makes clear to you His Signs that you may intellect"(Holy Quran 2:242)

"In Shia Islam, intellect is a key component of faith. Intellect allows us to understand the creation of God"(Aga Khan IV, July 23rd 2008, Lisbon, Portugal)

"......The Quran tells us that signs of Allah’s Sovereignty are found in the contemplation of His Creation - in the heavens and the earth, the night and the day, the clouds and the seas, the winds and the waters...."(Aga Khan IV, Kampala, Uganda, August 22 2007)

" Islam, but particularly Shia Islam, the role of the intellect is part of faith. That intellect is what seperates man from the rest of the physical world in which he lives.....This notion of the capacity of the human intellect to understand and to admire the creation of Allah will bring you happiness in your everyday lives. Of that I am certain"(Aga Khan IV, Dar-es-Salaam, Tanzania, August 17th 2007)

"Of the Abrahamic faiths, Islam is probably the one that places the greatest emphasis on knowledge. The purpose is to understand God's creation, and therefore it is a faith which is eminently logical. Islam is a faith of reason"(Aga Khan IV, Spiegel Magazine interview, Germany, Oct 9th 2006)

"In sum the process of creation can be said to take place at several levels. Ibda represents the initial level - one transcends history, the other creates it. The spiritual and material realms are not dichotomous, since in the Ismaili formulation, matter and spirit are united under a higher genus and each realm possesses its own hierarchy. Though they require linguistic and rational categories for definition, they represent elements of a whole, and a true understanding of God must also take account of His creation. Such a synthesis is crucial to how the human intellect eventually relates to creation and how it ultimately becomes the instrument for penetrating through history the mystery of the unknowable God implied in the formulation of tawhid."(Azim Nanji, Director, Institute of Ismaili Studies, London, U.K., 1998)

"Education has been important to my family for a long time. My forefathers founded al-Azhar University in Cairo some 1000 years ago, at the time of the Fatimid Caliphate in Egypt. Discovery of knowledge was seen by those founders as an embodiment of religious faith, and faith as reinforced by knowledge of workings of the Creator's physical world. The form of universities has changed over those 1000 years, but that reciprocity between faith and knowledge remains a source of strength"(Aga Khan IV, 27th May1994, Cambridge, Massachusets, U.S.A.)

"The Divine Intellect, Aql-i Kull, both transcends and informs the human intellect. It is this Intellect which enables man to strive towards two aims dictated by the faith: that he should reflect upon the environment Allah has given him and that he should know himself. It is the Light of the Intellect which distinguishes the complete human being from the human animal, and developing that intellect requires free inquiry. The man of faith, who fails to pursue intellectual search is likely to have only a limited comprehension of Allah's creation. Indeed, it is man's intellect that enables him to expand his vision of that creation"(Aga Khan IV, Aga Khan University Inauguration Speech, Karachi, Pakistan, November 11, 1985)

"The Holy Qu'ran's encouragement to study nature and the physical world around us gave the original impetus to scientific enquiry among Muslims. Exchanges of knowledge between institutions and nations and the widening of man's intellectual horizons are essentially Islamic concepts. The Faith urges freedom of intellectual enquiry and this freedom does not mean that knowledge will lose its spiritual dimension. That dimension is indeed itself a field for intellectual enquiry. I can not illustrate this interdependence of spiritual inspiration and learning better than by recounting a dialogue between Ibn Sina, the philosopher, and Abu Said Abu -Khyar, the Sufi mystic. Ibn Sina remarked, "Whatever I know, he sees". To which Abu Said replied," Whatever I see, he knows"."(Aga Khan IV, Aga Khan University Inauguration Speech, Karachi, Pakistan, November 11th 1985)

"In Islamic belief, knowledge is two-fold. There is that revealed through the Holy Prophet (s.a.s.) and that which man discovers by virtue of his own intellect. Nor do these two involve any contradiction, provided man remembers that his own mind is itself the creation of God. Without this humility, no balance is possible. With it, there are no barriers. Indeed, one strength of Islam has always lain in its belief that creation is not static but continuous, that through scientific and other endeavours, God has opened and continues to open new windows for us to see the marvels of His creation"(Aga Khan IV, Aga Khan University, 16 March 1983, Karachi, Pakistan)

"Our religious leadership must be acutely aware of secular trends, including those generated by this age of science and technology. Equally, our academic or secular elite must be deeply aware of Muslim history, of the scale and depth of leadership exercised by the Islamic empire of the past in all fields"(Aga Khan IV, 6th February 1970, Hyderabad, Pakistan)

"Islamic doctrine goes further than the other great religions, for it proclaims the presence of the soul, perhaps minute but nevertheless existing in an embryonic state, in all existence in matter, in animals, trees, and space itself. Every individual, every molecule, every atom has its own spiritual relationship with the All-Powerful Soul of God"(Memoirs of Aga Khan III, 1954)

"Islam is fundamentally in its very nature a natural religion. Throughout the Quran God's signs (Ayats) are referred to as the natural phenomenon, the law and order of the universe, the exactitudes and consequences of the relations between natural phenomenon in cause and effect. Over and over, the stars, sun, moon, earthquakes, fruits of the earth and trees are mentioned as the signs of divine power, divine law and divine order. Even in the Ayeh of Noor, divine is referred to as the natural phenomenon of light and even references are made to the fruit of the earth(Aga Khan III, April 4th 1952, Karachi, Pakistan)

"The God of the Quran is the One whose Ayats(Signs) are the Universe in which we live, move and have our being"(Aga Khan III, April 4th 1952, Karachi, Pakistan)

"Nature is the great daily book of God whose secrets must be found and used for the well-being of humanity"(Aga Khan III, Radio Pakistan, Karachi, Pakistan, February 19th 1950)

"In fact this world is a book in which you see inscribed the writings of God the Almighty"(Nasir Khusraw, 11th century Fatimid Ismaili cosmologist-philosopher-poet)

"O brother! You asked: What is the [meaning of] `alam [world] and what is that entity to which this name applies? How should we describe the world in its entirety? And how many worlds are there? Explain so that we may recognize. Know, O brother, that the name `alam is derived from [the word] `ilm(knowledge), because the traces of knowledge are evident in [all] parts of the physical world. Thus, we say that the very constitution (nihad) of the world is based on a profound wisdom"(Nasir Khusraw, 11th century Fatimid Ismaili cosmologist-philosopher-poet, from his book "Knowledge and Liberation")

“The physician considers [the bones] so that he may know a way of healing by setting them, but those with insight consider them so that through them they may draw conclusions about the majesty of Him who created and shaped [the bones]. What a difference between the two who consider!”(Abu Hamid Al-Ghazali, Muslim Theologian-Philosopher-Mystic, d1111CE)

"Tarkib' is composition as in the compounding of elements in the process of making more complex things, that is, of adding together two things to form a synthesis, a compound. Soul composes in the sense of 'tarkib'; it is the animating force that combines the physical elements of the natural universe into beings that move and act. Incorporating is an especially apt word in this instance. It means to turn something into a body, as in 'composing'. But it is actually the conversion of an intellectual object, a thought, into a physical thing. Soul acts by incorporating reason into physical objects, the natural matter of the universe and all the things composed of it"(Abu Yakub Al-Sijistani,10th century Fatimid Ismaili cosmologist, d971CE, from the book, 'Abu Yakub Al-Sijistani: Intellectual Missionary', by Paul Walker)

"One hour of contemplation on the works of the Creator is better than a thousand hours of prayer"(Prophet Muhammad, circa 632CE)

"All human beings, by their nature, desire to know."(Aristotle, The Metaphysics, circa 322BC)

24 December 2010

The Bacterial Flagellum – Truly An Engineering Marvel!

Jonathan M

A few weeks ago, I had the opportunity to attend a lecture by microbiologist Phillip Aldridge, of the University of Newcastle. The topic of his lecture was “The Regulation of Flagellar Assembly”. Being an ID proponent, I had a natural interest in what Aldridge was going to say, and I had been looking forward to the event for some time. I was already familiar to a degree with several of the key mechanisms and regulation of flagellar biosynthesis. Nonetheless, the lecture succeeded in re-kindling my passion for biology, and inspired me to do some in-depth research on my own with regards the workings of this engineering marvel.

I must confess that I was blown away. If one thought that the functional-specificity of arrangement with respect to the flagellum’s key components may well provide adequate grounds for a design inference, the mechanisms of flagellar construction take this intuition to a whole new level. So mesmerized I was by the motor’s intrinsic beauty and elegance, that I decided to provide a sketch overview of this amazing process for the benefit of readers of this blog. Of course, there are variations in the flagellum’s overall construct from species to species. The archetypical flagellum, however, is probably that of the closely related species, Escherichia coli, Salmonella enterica, and Salmonella typhimurium. It is this that I want to primarily focus on.

An overview of the flagellum’s construct

As with a typical man-made motor, the unit which provides rotary motion (which is embedded in the inner cell membrane) revolves within a stationary component (stator). The rotor is a tube-like structure which extends from within the cell, through the membrane, to the outside, where the flagellar filament (which serves as a propeller) is attached via a flexible joint.

The flagellum consists of a basal body (which is embedded in the cell wall) and two axial structures, the book and filament. The basal body is made up of the MS ring, rod, and L- and P- rings. Parts of the axial structure are exported from the cell by the type III secretion system (T3SS). The T3SS is composed of several proteins from the MS ring and a peripheral hexameric ATPase FliI that drives the export process.

The motor is anchored in the cytoplasmic membrane and cell wall. The motor consists of a central rod that passes through a series of rings. In gram-negative bacteria, an outer ring (the L ring) is anchored in the lipopolysaccharide layer. Another ring (the P ring) is anchored in the peptidoglycan layer of the cell wall. A third set of rings (the MS and C rings) are respectively found within the cytoplasmic membrane and the cytoplasm. Gram positive bacteria, by contrast, do not have an outer membrane, and thus only possess the inner pair of rings. A series of proteins, called Mot proteins, surround the inner ring and are anchored in the cytoplasmic membrane. The structures of the L, P, C and MS rings, in combination with the central rod, are collectively referred to as the “basal body”. In addition, a class of proteins called the Fli proteins function as a ‘switch’ for the reversal of motor rotation, which is plugged into an elegant signal transduction mechanism for receiving feedback from the environment — I will discuss this further in due course.

The flagellar filament (propeller) is constructed from subunits of the polypeptide flagellin. There is a wider region at the base of the filament (the ‘hook region’), which connects the filament to the motor component of the base. These subunits are arranged in a tight helical structure to produce a stiff hollow rod. Each subunit contributes to a small propeller blade to the outside of the structure.

The driving force behind the flagellum’s rotary motion comes from a proton motive force, to which I shall return in due course.

Chemotaxis and Signal Transduction

Chemotaxis refers to the means by which bacteria direct their movements in response to environmental chemical signals. This mechanism allows the bacteria to locate food by swimming towards the highest concentration of food molecules (such as glucose). The default direction of flagellar rotation is anticlockwise. This directionality of rotation can be reversed by virtue of a classic two-component signal transduction circuit (discussed below). When the direction of rotation is switched to clockwise, the flagellar bundle breaks apart, and the bacterium physically “tumbles” in place. The overall movement of a bacteria can be described in terms of a series of alternating swimming and ‘tumbling’ phases. When the bacterial cell ‘tumbles’, it re-orientates itself. By repeatedly evaluating its course, the bacterium is able to find favourable locations with the highest concentration of attractant (possibly a food source).

A two-component signal transduction circuit essentially involves a histidine protein kinase that catalyses the transfer of phosphoryl groups from ATP to one of its own histidine residues, and a response regulator that catalyses the transfer of phosphoryl groups from the histidine kinase to its own aspartate residue. In the chemotaxis system, the histidine kinase, CheA, associates with chemoreceptors (a distinct class of transmembrane receptor proteins) which interact with chemicals in the surrounding environment. The switch in flagellar direction is induced by CheY, which interacts with a flagellar switching protein, FliM, inducing the change from counter-clockwise to clockwise rotation.

The chemoreceptors which form a part of the chemotaxis system are Tsr, Tar, Tap, Trg, and Aer. In combination with CheA and CheW, they act to form receptor-signalling complexes that function to integrate sensory information to regulate CheA autophosphorylation, hence controlling its activity. This, in turn, controls the phosphorylation of the response regulator (CheY). Ordinarily, CheY is reversibly bound to CheA. When it becomes phosphorylated, however, it dissociates with CheA and quickly diffuses to the flagellar motors. The CheY then acts as an allosteric regulator to promote the clockwise rotation of the flagellar bundle, which results in the ‘tumbling’ motion. An attractant stimulus interacts with the chemoreceptors, inhibiting the activity of the CheA kinase. This results in a decrease in phosphorylated CheY. Thus, the propensity of the bacterium to re-orient itself by “tumbling” is decreased.

When CheB is activated by CheA, it acts as a methylesterase, and removes methyl groups from glutamate residues on the cytosolic side of the receptor. It acts antagonistically with CheR, which adds methyl residues to the same glutamate residues. If the attractant’s concentration is high, the level of CheA phosphorylation (and, hence, CheY and CheB phosphorylation) stays relatively low. The bacterium thus swims relatively smoothly. Because the phosphorylated CheB is not available to demethylate, the level of methylation of the chemoreceptors increases.

Methylation, however, inhibits the chemoreceptors from responding to the attractant. Thus, the level of phosphorylated CheA and CheB increases and the bacterial cell tumbles, regardless of whether the level of the attractant is high. The availability of CheB-P now means that the chemoreceptors can be demethylated, and the receptors become available again to respond to attractants. In the case of repellants, the system works in exactly the opposite way — whereby more methylated chemoreceptors respond most and the lesser methylated chemoreceptors respond least!

A diagrammatic illustration of this system is to be found in the figure below:

Proton Motive Force

A proton motive force drives the rotation of the bacterial flagella. Electrochemical energy is converted into torque via an interaction between the stator and the rotor. Torque is then transmitted from the C-ring by the MS ring to the rod and then to the hook region. From there it is transferred to the propeller (filament). When the filament rotates, the torque is converted into thrust, allowing the bacterial cell to move.

The rotary engine is located at the flagellum’s anchor point on the inner membrane. A flow of protons across the membrane, resulting from a concentration gradient, drives the motor. Protons are transferred across the membrane by the rotor, and the rotor is turned in the process. The speed at which the motor turns can be increased or decreased as a direct result of changes in the strength of the proton motive force.

But it only gets better. An interesting 2008 study by Blair et al. demonstrated that an operon required for the biosynthesis of the biofilm matrix in Bacillus subtilis also encodes a molecular clutch, called EpsE. When associated with a flagellar basal body, EpsE disables the biological motor by disengaging the drive train from the power source! Those hungry for further reading are invited to read Howard Berg’s 1995 paper on Torque generation in the bacterial flagellar rotory motor.

Flagellar Assembly

The processes of flagellar assembly, and its regulation, is my personal unparalleled favourite area of study in the discipline of microbiology. For an informative documentary on the assembly of this nanomachine, I highly recommend viewing this video.

The self-assembly of the bacterial flagellum requires the co-ordinated expression of more than 60 gene products (which encode for both structural and regulatory proteins). Bacteria are intriguingly able to detect the stage of progress with respect to flagellar assembly, and subsequently use this information in the co-ordination of gene expression.

The mechanisms of flagellar assembly are so complex and so sophisticated that justice can hardly be done to them here. Interested readers who desire a fuller discussion of these mechanisms are advised to read a 2002 paper, authored by Phillip Aldrige and Kelly T Hughes, appearing in Current Opinion in Microbiology. In addition, a technical book, Pili and Flagella: Current Research and Future Trends features a detailed discussion of this topic. The book is typically priced at £150/$310, but a significant portion of the book may be accessed via Google Books. The book discusses studies of flagellar mutants that fail to complete their flagella. In so doing, they discern which proteins are absolutely indispensible to flagellar assembly, and which are not — something any ID proponent should be interested in.

Flagellar assembly begins in the cytoplasmic membrane, progresses through the periplasmic space, and finally extends outside the cell. As previously alluded to, the flagellar apparatus consists of two main parts: the secretion system and the axial structure. The main components of the axial structure are FlgG for the rod, FlgE for the hook, and FliC for the filament. All of these assemble with the help of a cap protein (FlgJ, FlgD and FliD respectively). Of those, only FliD remains at the filament’s tip — the other two are not present in the finished product. Other components of the axial structure (FlgB, FlgC and FlgF) connect the rod and MS ring complex. The hook and the filament are connected by FlgK and FlgL

The MS ring complex is the structural foundation of the apparatus. When the C ring and C rod attach to the M ring (at its cytoplasmic surface), the complex begins to secrete flagellar proteins.
With the assistance of a cap protein (FlgJ), the rod structure is constructed through the peptidoglycan layer. But its growth is terminated when it reaches the outer membrane (which represents a physical barrier such that it cannot pass through unaided). But not to worry! The outer ring complex cuts a hole in the membrane. The hook then begins to grow beneath the FlgD scaffold until it reaches approximately 55nm in length. When the hook reaches this critical length, the substrates which are being secreted switches from the rod-hook mode to flagellin mode. FlgD is then replaced by HAPs (hook associated proteins) and the filament continues to grow — note that this can only take place in the presence of FliD (the cap protein), otherwise the flagellin monomers are lost.

The Regulation of Flagellar Assembly

The energy costs of assembling flagella make these nanomachines expensive systems to run. This entails the necessitude for systems and mechanisms to regulate the co-ordination of assembly. The flagellar system of Salmonella (the paradigm organism for this system) possesses three classes of flagellar promoters which are organised into a transcriptional hierarchy. The Class I promoter drives the expression of the enteric master regulator, FlhD4C2. In association with the sigma factor, σ70, this master regulator turns on the Class II promoters which are responsible for the gene expression of the hook-basal-body subunits and its regulators, including σ28, and its anti-sigma factor, FlgM. σ28 is required for the activation of the Class III promoters. This class of promoters is responsible for the expression of flagellin monomers, the chemotaxis system and the motorforce generators. Interaction with its anti-sigma factor (FlgM), thus keeping it away from the RNA polymerase holoenzyme complex, inhibits the activation of σ28 prior to the completion of the Hook Basal Body. When the Hook Basal Body is finished, the FlgM is secreted through the flagellar structure via the Type III Secretion Apparatus substrate specificity switch. This means that the Class III promoters can now be activated by their sigma factor (σ28) because its anti-sigma factor (which represses its action) has been removed. This allows the flagellum to be completed.

This brief overview of flagellar assembly has barely scratched the surface of this intricate process. Nothing will please me more than if, after having read this description, at least a few readers reach for the relevant literature to find out more.

The Bacterial Flagellum — A Paradigm for Design?

In writing the above descriptions, I hoped to treat the educated layperson to a flavour of the magnificence of this rotary motor. As previously stated, nothing will bring me more satisfaction than having provided someone with the inspiration to pursue further information. The question does arise, however, as to whether this system bears the hallmarks of design — or is it, as so many biologists maintain, merely the product of blind, purposeless and impersonal natural processes of chance and necessity? To me, the answer is very clear. This system certainly appears to be designed technology. It certainly appears to bear the hallmarks of design logic. And it does seem that the burden of proof must, at this present time, lie with he who rejects this proposition — that is, he who ascribes this system to blind natural processes. The handwaving gestures of Kenneth Miller, who asserts that the Darwinian explanation for this system is more appropriate in light of the presence of the Type III Secretion System, can no longer be considered a satisfactory rebuttal to the design postulate. Such gestures may, at first glance, appear convincing to the uninitiated, unfamiliar with the complexities of these systems. But when one examines the molecular structure and orchestration of these systems, one loses satisfaction with the vague discussions of protein sequence homologies. Kathryn Applegate and Ard Loius seem to be under the impression that this assembly process parallels the processes of Darwinian evolution — I will allow readers to judge for themselves on that one.

As has been pointed out by Stephen Meyer and Scott Minnich, there is an even more fundamental challenge to the standard Darwinian line with regards this system. Firstly, the phylogenetic evidence is strongly suggestive that the T3SS is an evolutionary biproduct of the flagellar system, rather than the other way round (see, for instance, here). Secondly, if flagellum biosynthesis were to be expressed simultaneously with the Yop T3SS, flagellin monomers would likely be exported out the needle-like structure as well as the flagellar basal body, potentially limiting the efficiency of both systems. Meyer and Minnich argue in their paper that the potential for cross-recognition between Type III exported proteins in the same cell explains why the segregation of these systems by specific environmental cues is necessary. Expression of a flagellum under host conditions would result in a loss of polarised secretion of Yop proteins into the cells of the host. Flagellin is also a potent cytokine inducer – display of flagellin to the macrophages by direction injection via the Ysc secretin would strongly countermand the Yersinia’s anti-inflammatory strategy.

Easy Nash

In Shia Islam, intellect is a key component of faith. Intellect allows us to understand the creation of God: Aga Khan IV(2008)
The Qur'an itself repeatedly recommends Muslims to become better educated in order better to understand God's creation: Aga Khan IV(2007)
The Quran tells us that signs of Allah's Sovereignty are found in the contemplation of His Creation: Aga Khan IV(2007)
This notion of the capacity of the human intellect to understand and to admire the creation of Allah will bring you happiness in your everyday lives: Aga Khan IV(2007)
Islam, eminently logical, placing the greatest emphasis on knowledge, purports to understand God's creation: Aga Khan IV(2006)
The Holy Qu'ran's encouragement to study nature and the physical world around us gave the original impetus to scientific enquiry among Muslims: Aga Khan IV(1985)
The first and only thing created by God was the Intellect(Aql): Prophet Muhammad(circa 632CE)