What’s in a name?

What’s in a name? That which we call lactate dehydrogenase by any other name would still convert lactate to pyruvate; so would EC 2.4.1.11 (glycogen synthase) were it not EC 2.4.1.11 call’d…..

I think many people share these sentiments when it comes to the naming of enzymes. I really don’t think that it matters whether the enzyme is coded and classified or just named for its function. For all that matters I have no problem calling the enzyme in each reaction as enzyme 1 or glucose breaker.

But alas, if life were so easy then there would be a lot of chaos in the scientific world since what I will be calling hexokinase in glycolysis, some person in Australia might be calling it Bob’s enzyme. Therefore for sake of convenience and standardization, there has been a system created to assign names to all the known enzymes.

Now, pay close attention cause I will attempt to breakdown this whole nomenclature business with enzymes. Enzyme nomenclature is assigned by two names: recommended name and systematic name.

Recommended name is the commonly referred to names or most recognized names based on their function, their substrate or just by who discovered them/random naming. You might see the problems here with lack of uniformity and a bit of confusion in the naming. Examples of this type of naming may include lipases (enzymes involved in the metabolism of lipids), pyruvate decarboxylase (removes the carboxylic acid group to form carbon dioxide) or even trypsin (hydrolysis of peptide bonds).

Systematic name was developed by the International Union of Biochemistry and Molecular Biology (IUBMB). This group of biochemists came together in a joint committee and decided that it was time to standardize and codify enzymes. The main purpose was to facilitate better communication of all biochemical information and research by having a standard generally understood nomenclature and description of enzymes. In 1955, they devised a system of nomenclature that divided all enzymes into 6 major classes and then subdivisions within each class. These names were based completely on the reaction to which the enzyme catalyses and includes all the substrates involved as well. This provided a comprehensive and informative method to which enzymes may be named. This name was also codified for specificity and reference. However, this is quite a mouthful to use and still enzymes are usually referred to by their recommended names.

Below is an illustration of the 6 major classes of the systematic nomenclature.

Before I begin, you might be wondering well, how am I going to remember these 6 classes? Well, I have devised a mnemonic, which is slightly rude, but don’t judge me, because I think that the ruder mnemonics are, the more they are remembered.

Okay, so there are 6 major classes:

  1. Oxidoreductases
  2. Transferases
  3. Hydrolases
  4. Lyases
  5. Isomerases
  6. Ligases

So, taking the first letter from each class we have:

Only

Two

Happy

Lovers

Indulge in

Lovemaking

All right, so that may aid in remembering the order of the classes, but unfortunately, we still have to learn the details of each class. Also knowing the class, will aid in understanding the coded name for the enzyme. This coded name is known as the ENZYME COMMISSION NUMBER (EC number). The EC number can be broken-down as follows:

Using the example of glycogen synthase

EC 2.4.1.11

2: represents the class to which it belongs; transferases

4: represents the subclass that transfers carbon-groups to the glycogen

1: sub-subclass

11: serial number

This gives insight into the specific action of the enzyme.

Right, now that all that is out of the way, let us now focus on each class, where I will give a brief description of each and follow with an example.

  1. Oxidoreductases: as the name implies these enzymes catalyse oxidation- reduction reactions (redox reactions). Example: alcohol dehydrogenase. This enzyme facilitates the conversion of alcohols to aldehydes by reduction of NAD  to NADH. Its systematic name is alcohol: NAD+ oxidoreductase. Its EC number is EC 1.1.1.1
  2. Transferases: these enzymes catalyse the transfer of carbon, nitrogen or phosphate groups in a reaction. Example: tRNA (cytosine-5-)-methyltransferase. This enzyme facilitates the conversion of S-adenosyl-L-methionine to S-adenosyl-L-homocysteine by transferring a methyl group from methionine. Its systematic name is S-adenosyl-L-methionine: tRNA (cytosine-5-)-methyltransferase. Its EC number is EC 2.1.1.29
  3. Hydrolases: these catalyse the cleavage of bonds by the addition of water molecules. Example: acetylcholinesterase. This enzyme facilitates the degradation of the neurotransmitter acetylcholine to acetate, choline and hydrogen ions by addition of water molecule. Its systematic name is acetylcholine acetylhydrolase. Its EC number is EC 3.1.1.7
  4. Lyases: these catalyse the cleavage of C-C, C-S and certain C-N bonds by other means than hydrolysis and oxidation. Example: carnitine decarboxylase. This enzyme facilitates the degradation of carnitine to 2-metylcholine by the cleavage of the C-C bonds to form carbon dioxide. Its systematic name is carnitine carboxy-lyase (2-methylcholine forming). Its EC number is EC 4.1.1.42
  5. Isomerases: these catalyse the racemization (conversion of an enantiomer to another enantiomer) of optical or geometric isomers. Example: ribose-5-phosphate isomerase. This facilitates the conversion of D-ribose 5-phosphate to D-ribulose 5 phosphate. This involves the conversion of the pentose sugar from the aldose form to the ketose form. Its systematic name is D-ribose-5-phosphate aldose-ketose-isomerase. The EC number is EC 5.3.1.6
  6. Ligases: these catalyse the formation of bonds between carbon atoms and O, S, N atoms that are coupled with the hydrolysis of high-energy phosphate groups. Example: succinyl-CoA synthetase (ADP forming). This enzyme facilitates the conversion of succinate to succinyl-CoA and driving the formation of ADP by the cleavage of the phosphate group from ATP. The systematic name is succinate-CoA ligase (ADP-forming). The EC number is EC 6.2.1.5

Well, well, well. That was a lot to take in but read it through and understand that basics of the classes and the EC classification. I really hope that this review helps and did not confuse you at all.

I believe that you can now realize the importance of the nomenclature of enzymes. Peace out for now people and enjoy the biochem.

 

References

 

Protein and amino acids questions

test time

Well you know the deal, answer the questions below if you are ever so gracious. Please and thank you. Hit me up in the comment section to answer them. I will give feedback accordingly.

Exam questions

1. Which is amino acid residue is NOT generally found in the alpha-helix structure of proteins?

a)    Cysteine

b)   Arginine

c)    Glycine

d)   Lysine

e)    Glutamate

 

2. Which group of amino acids are essential amino acids?

a)    Methionine, glutamate, tyrosine

b)   Cysteine, glycine, alanine

c)    Arginine, aspartate, glutamine

d)   Valine, phenylalanine, lysine

e)    Tryptophan, proline, isoleucine

 

3. What is the reaction that forms a disulphide bridge?

a)    Reduction

b)   Oxidation

c)    Hydrolysis

d)   Phosphorylation

e)    Condensation

 

4. Select the correct multiple choice answer using ONE of the keys; A, B, C, D or E as follows:

a)    1, 2 and 3 are correct

b)   1 and 3 are correct

c)    2 and 4 are correct

d)   only 4 is correct

e)    all are correct

Which of the following is/are characteristic of collagen?

1. Possess proline and glycine amino acid residues

2. Requires ascorbic acid during formation/synthesis

3. Forms a triple helical structure

4. Is a globular protein

 

5. Which type of bonding is predominantly involved in the formation of beta-pleated sheet structure of proteins?

a)    Hydrophobic interactions

b)   Ionic bonding

c)    Covalent bonding

d)   Hydrogen bonding

e)    Metallic bonding

 

6. Select the correct multiple choice answer using ONE of the keys; A, B, C, D or E as follows:

a)    1, 2 and 3 are correct

b)   1 and 3 are correct

c)    2 and 4 are correct

d)   only 4 is correct

e)    all are correct

Which of the following statements accurately describes the Anfinsen experiment?

1. The main purpose of the experiment was to illustrate co-operative binding of the quaternary structure of proteins
2. A kosmotrope was used to denature the hydrophobic bonds present in the 3-D structure of RNase A
3. The biological activity of RNase A remained intact after the disulphide bridges were broken
4. The experiment supported the idea that all the information about protein folding was present in the amino acid sequence of the primary structure of RNase A

exam time

Reflection of the week 5

 Hey Biochem people, so week 5 has come and gone and this week was Amino acids and proteins. We are now getting into the real ‘meat’ of this biochem thing.

Amino acids and proteins are THE most important biomolecule in my opinion. These molecules are found in just about every nook and cranny of the body. As you may realize that we, humans are nothing without these little molecules.

 I enjoyed this week of lectures since it is the topics that lay the foundation to just about every other biochemical reaction in the body. Amino acids and proteins for that matter are critical to the efficient and effective functioning of the body.

 

Proteins act as the gladiators of our body fighting and defending intruders such as antigens that infiltrate our blood and other bodily fluids and try to suppress our immune system.

These invaders may be microbial villains like bacteria, viruses and even foreign objects.

 

These knights in shining armour are none other than…

ANTIBODIES. Antibodies work with our armies of immune cells such as white blood cells to suppress the enemy and keep the immune system functioning at optimal level.

 

 

Proteins also aid in the moving and shaking of cells and the body by extension….literally!

Contractile proteins such as myosin and actin interact with each other to facilitate movement via contraction of these molecules. This is observed within the cell to enable cytoplasmic streaming as well as in myocytes (muscles) that facilitate movement of muscular tissue. Therefore, we could not shake our moneymakers without proteins.

 

Proteins make up the working force of the body. They are there diligently labouring to speed up or facilitate the biochemical reactions of our body. These catalysts ensure that all the requirements for metabolism and general functioning are carried out. And you must guess what this protein is… ENZYMES. These things are literally everywhere and in everything… like a macco. They aid in digestion, energy production and degradation, drug metabolism, clean up and recycling.

 

Now, proteins may also be the traffic police and mail carriers of the body. They can transmit messages as well as co-ordinate different bodily processes. These molecules are essential to managing certain reactions. Hormones such as insulin produced by the beta cell of the islet of Langerhans in the pancreas crucial to the regulation of glucose intake and metabolism, somatotropin aka a growth hormone is critical to stimulation of protein production such as creatinine to make muscles.

collagen

Proteins also form the backbone to many structures of the body including skin, bone, organs and ligaments and various appendages like hair and nails. These fibrous, insoluble structures provide support to numerous structures. Structural proteins include collagen, keratin and elastin. These proteins are found in hair and connective tissue. Therefore, being able to whip your hair back and forth or jumping up and down without your joints giving way and crumpling like a puppet is all thanks to proteins.

 

 

Proteins may also be the big trucks of the body acting as carrier molecules that transport important molecules throughout the cell as well as the body to the target destination to be used.  The famous ones such as Haemoglobin that transports just about the most important molecule, oxygen! to cells as well as cytochromes which are critical to electron transport in ETC in the inner mitochondria.

electron-transport

 

And this is just a few of the many, many functions of proteins in the body. Hence, I believe that proteins are VERY GOOD THINGS!!!

 Overall, I am excited for the rest of the term to delve into the exciting path of this Biochem course.

 Reference

About.com Biology. 2013. “Protein Function by Regina Bailey.” Accessed on March 14, 2013. http://biology.about.com/od/molecularbiology/a/aa101904a.htm

Reflection of the week 6

 Alright people, we are half way there and some might be just zooming through,

while others, like me, are already feeling a burnout.

Nevertheless, we need to hang in there and keep the faith…. We are going to come out of it for the better.

So smile and keep telling yourself those positive lies.

 

Now on to business. This week was all about enzymes and tutorials. I again, cannot emphasize the importance of tutorials and quizzes since it really applies and tests your understanding of key concepts in the lectures. I will speak in depth about our little “cell workers” in subsequent postings since enzymes will form the basis onto which all other topics will build on.

 

However, before that, I want to speak about two things that were brought up in quiz and tutorials on amino acids and proteins. Some might have already guessed what I am talking about and that is the Anfinsen Experiment and the five factors that affect the alpha helix secondary structure of proteins.

Anfinsen experiment

 

Dr C Anfinsen was a biochemist from Pennsylvania who won the Nobel Prize in 1972 for his work in protein folding. Specifically, he proposed the idea that the information that determines the tertiary structure of proteins resides in the amino acid sequence and composition found in the primary structure. Through his investigations, he was able to show the reversible denaturation of proteins.

But exactly what did he do? And why is this so important to our course?

 

 

 

 

Well here is the 411 on what you need to know about this experiment.

First things first let us name the players:

Ribonuclease A enzyme: small, stable and easily purified. And its function is as the name suggests it, the enzyme that degrades RNA. Role in the experiment: it is the enzyme that is being denatured and reverse denatured. It is biologically active in the folded tertiary structure

Urea: This substance is member of a group of substances that you should never forget… A CHAOTROPE!!!!   Now a chaotrope is a substance that denatures proteins by disrupting the hydrophobic interactions responsible for maintaining the tertiary structure of the protein. It does this by destabilizing the hydrophobic interactions and cause unfolding of the protein. However, the disulphide bonds, if present, are a very strong bond and therefore will not be affected by this agent. And it is because of the disulphide bond, there is not complete unfolding of the protein during denaturing.

v

  Β-mecaptoethanol is a reducing agent that reduces the disulphide bonds present in the structure. This substance reacts on the disulphide bond of the cystine molecule reducing it to two cysteine molecules that possess sulphur hydryl groups. This breaking of the disulphide bond leads to unfolding and hence loss of biological activity of the enzyme. However, the primary linear structure of the protein remains intact.

Dialysis tubing: this is a partially permeable tubing that contains very small pores that allows for diffusion of molecules from a state of high concentration to a state of low concentration. Visualize it as a penna-cool pack with lots of tiny pinprick holes throughout the body.

 

Okay, so now that we have laid out the key components of this experiment, let us get down to it.

1.   Ribonuclease A was treated with the chaotrope, urea, which caused the unfolding due to disruption of the hydrophobic interactions, however, complete denaturation did not occur and the ribonuclease A retained low, but still present, biological activity and degraded the RNA.

But why???????

Oxidized ribonuclease with the disulphide bonds

Oxidized ribonuclease with the disulphide bonds

And the simplest answer, because of the presence of the disulphide bonds. This strong bond found in cystine, is formed by the oxidation of two cysteine residues. Therefore, something is needed to break this bond if some unfolding is to occur.

Reduced Ribonuclease with disuphide bridges hydrolyzed

Reduced Ribonuclease with disuphide bridges hydrolyzed

2.   Beta-mecaptoethanol was added to the solution of urea and ribonuclease A. The mecaptoethanol reduces the cystine residue by breaking the disulphide bond and form two cysteine residues. The hydrolysis of the disulphide bond causes the complete unfolding of the tertiary structure of the enzyme and hence loss of the biological activity. However, the primary structure of amino acids linked together in a chain by peptide bonds will not be harmed.
3.   Well now that ribonuclease is denatured, the next step is to prove the theory that the information necessary to form the tertiary structure is encoded in the amino acid sequence of the primary structure. To perform this, the solution containing the enzyme, mecaptoethanol and urea is added to the dialysis tubing. The tiny pores present in the tubing allow for the diffusion of mecaptoethanol and urea since thy are tiny enough to pass through the pores. But it leaves the ribonuclease in the tubing since it is too large to pass through the pores.

4.   In the absence of the urea and mecaptoethanol, the hydrophobic interactions may reform and there is oxidation of the two-cysteine residues to reform the disulphide bonds of cystine. This means that in the absence of the chaotrope and reducing agent, there is refolding of the protein and regaining of the biological function.

Native biological active ribonuclease

Native biological active ribonuclease

And there you have it. Now that was not that bad and hopefully it is easy to understand and not too difficult to read.

Well in prepping this little explanation, I came across a blog by Laurence Moran, who conveniently co-authored one of the biochem text that I use. In his blog, he explains this experiment and the important pointers to note. Now it is not at all in depth, but I feel that it summarizes the essential points to take away and reinforces those concepts of reducing agents, chaotropes, disulphide bonds and protein folding. So I am placing the link to the blog below. Take a browse and see if you understand. He has some diagrams as well which may aid in understanding. I felt this was well explained and good little revision to consolidate all the info already presented on this experiment.

Sandwalk. Strolling with a sceptical biochemist. The Anfinsen Experiment in Protein folding by Laurence A. Moran.

http://sandwalk.blogspot.com/2007/02/anfinsen-experiment-in-protein-folding.html

I also came across a YouTube video on the Anfinsen experiment by Suman Bhattacharjee. He also explains this experiment in a comprehensive yet simple manner that gives good insight into the fundamentals and the important points to note. However, as you would infer by his name, he is Indian and therefore has quite a thick Indian accent that makes it difficult at some points to full understand what he is trying to say. However, this is minor since the material he is teaching is very valuable and well explained. The added fun of this is that it is a video and therefore little reading needs to take place. This is a benefit that I do not under estimate. Below is the information to check it out yourself.

Anfinsen experiment by Suman Bhattacharjee

http://www.youtube.com/watch?v=zaaAolMw25A

Five factors affecting the alpha helix of proteins.

 

Now we all know, I hope, what the alpha helix is and its structure. Well let us just summarize by saying that the alpha helix is more stable secondary structure of proteins that is formed by the hydrogen bonding of the carbonyl group of the peptide bond of amino acids in the primary structure to the hydrogen on the amino group of the peptide bond 4 amino acid residues away. This causes a helical arrangement of the amino acids producing turns that are 0.54 nm in distance and contain approximately 3.6 amino acid residues. It is the most stable secondary structure since it possesses maximal amount of hydrogen bonding where almost all the amino acids except for those residues present at the ends of the helix, are involved in hydrogen bonding.

So we are hopefully all on the same page now and let us go through the five main factors that affect the alpha helix.

1.   Proline and glycine
 

Proline is a non-polar aliphatic amino acid that contains a ringed structure in its R group. As can be seen the side chain consists of CH2-CH2-CH2. This ringed structure is bonded to both the alpha carbon and alpha amino group. This causes two problems. Firstly, the bonding to the alpha amino group causes the rigidity of the alpha carbon-N bond on the structure. This prevents the rotation of this bond and therefore will cause the inability to bend and conform to undergo hydrogen bonding in the helix. This destabilizes the helical structure and causes a kink in the alpha helix structure. Proline also lacks a hydrogen atom in its amino group (NH2 instead of NH3) since it is bonded to the ring of methyl groups and therefore is lacking the hydrogen bond necessary to perform hydrogen bonding. This will also destabilize the helix. Therefore, proline is rarely found in the alpha helix.
 

Glycine is the smallest known amino acid and is also classed under the non-polar aliphatic amino acid. It contains a H-atom as the R- group. This causes high conformational flexibility and therefore will cause destabilization of the alpha helix. This is why glycine may not be found in alpha helix.

 

2.   Long segments of positively or negatively charged amino acid residues
 

Within the helix, if there are long segments of positively charged amino acids such as lysine, histidine and arginine, there is a repulsion of forces causing a destabilization of the helix. Similar actions will occur if there are long segments of negatively charged amino acids such as glutamate and aspartate.

3.   Steric hindrance/interference
 
As the hydrogen bonding causes the formation of the helix, the R-groups point outwards away from the helix. In some circumstances, the bulky R-groups may come into close proximity of each other and this bulky R-group may prevent proper hydrogen bonding and disrupt the stability of the helix. Aromatic amino acids such as phenylalanine, tyrosine and tryptophan are examples of amino acids that have bulky R-groups that prevent proper hydrogen bonding in the helix.
 
4.   The interaction of positive and negative amino acid side chains.
 
As the helix turns, positively charged amino acids such as lysine, arginine and histidine may come into close proximity of the side chains of negatively charged amino acids such as aspartate and glutamate. If a positively charged amino acid side chain such as lysine comes into close proximity to the side chain of negatively charged glutamate, there is the formation of an ion pair or salt bridge. This causes the stabilization of the alpha helix.
 
5.   The position of amino acids.
 
This is probably the most important factor of all. It deals with the position of certain amino acids that will facilitate the increase in stabilization of the alpha helix. But before we discuss any amino acids, let us take a step back and talk a little about the alpha helix. First off, the hydrogen bonds that are formed to create this secondary structure run parallel to the imaginary upright axis of the alpha helix. Every hydrogen bonding causes the formation of a dipole. But you must be asking yourself, but what is a dipole?
 

Well, a dipole is a pair of equal but opposite electric charges or magnetic poles that are separated by a small distance. This means that each hydrogen bonding causes the formation of mini-dipoles. Since these are distributed one on top of each other in the helix. This causes all the micro-dipoles formed to exert a large dipole on the entire helix. This means that all the small dipoles are consolidated to form a macro dipole. This macro dipole of the entire helix means that one end of the helix exerts a slightly positive charge while the other exerts a slightly negative charge. This alludes to the fact that the N-terminal end or the end that possesses the free amino group, will be slightly positively charged while the C-terminal end or the end possessing the free carboxylic group will be slightly negatively charged.
 

Alright, now to the amino acids and their role in all this. Therefore, negatively charged amino acids such as glutamate and aspartate are found on near the N-terminal end since it will be attracted to the slightly positive charge that leads to the formation of ion pairs and this is cause an interaction that will stabilize the helical structure. Positively charged amino acids such as lysine, arginine and histidine may be found near the C-terminal end since the opposite charge causes an interaction and formation of ionic bonds that will further stabilize the helix. Whereas negatively charged amino acids, glutamate and aspartate will be positioned near the N-terminal end since there is attraction of the positive charge will also cause stabilization of the helix.

 

Well there you have it folks, a little summary of two very important concepts. I trust that everything is easily understandable. Hit me up on the comment section if you see anything that may be wrong or any clarifications you want to make. Bye for now and keep trudging along.

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 Reference

  • Ribonuclease A: September 2008 Molecule of the Month by David Goodsell. 2013

http://www.rcsb.org/pdb/101/motm.do?momID=105

  • Nobelprize.org: The Nobel Prize in Chemistry 1972. 2003

http://www.nobelprize.org/nobel_prizes/chemistry/laureates/1972/anfinsen-bio.html