Why I Am Interested in Structural Biology
Juan Guajardo

Introduction

The purpose of this experiment was to find out the best conditions for the ideal crystal growth of the enzyme lysozyme, the abundant protein found in our tears and in hens eggs. The crystals will then be used to study other less well-known proteins, and someday be able to produce better medicines to prevent certain illnesses like diabetes, HIV Reverse Transcriptase, Human Rhino Virus, etc. Because of gravity the crystals do not grow very big on Earth and cannot be examined with an X-ray beam to get a diffraction pattern and obtain the structure of the protein. Lysozyme crystals are taken to space because the microgravity environment provides a convection and sedimentation free environment resulting in bigger and better quality crystals suitable for examination. The Central Dogma of Molecular Biology forms the backbone of molecular biology and has four major stages: replication, transcription, procession, and translation. First the DNA replicates its data in a procedure that includes many enzymes. Then, the DNA codes for the production of messenger RNA (mRNA) during transcription. Next the mRNA is processed and moves from the nucleus to the cytoplasm, the colloidal substance surrounding the nucleus and other structures in living cells. Finally, the mRNA carries coded data to the ribosomes. Then the ribosomes translate this data and use it for protein synthesis.

Hypothesis

My hypothesis for the protein crystal experiment was that that sample six (.50g of NaCl) would work best because its concentration was not too much and not too little. I thought that sample eight would have large amount of crystals but would probably be too small. I predicted that sample one would have very few and small crystals. The predictions below are for the number and the size of the crystals in the first day and the second day.

SAMPLE # 1 2 3 4 5 6 7 8
Day1prediction 10 12 20 25 28 31 43 56
Day2prediction 12 18 25 24 15 50 75 56
Day1prediction .1mm .2mm .3mm .4mm .5mm .5mm .3mm .2mm
Day2prediction .1mm .1mm .4mm .5mm .5mm .4mm .4mm .3mm

Procedure

The materials the team used were: an Erlenmeyer flask, eight small test tubes, a funnel, a gelatin capsule, a snap-cap tube, a dropper pipette, a 15mL measuring tube, acetate buffer, lysozyme, and salt.

After getting the materials, our team started to conduct the experiment by first numbering the tubes one through eight. Then we measured out 10mL of the buffer into the 15mL conical tube, and then poured the buffer solution into the Erlenmeyer flask. Next, our team placed the funnel into the flask and carefully one member poured the lysozyme into the flask. After waiting five minutes for the lysozyme to dissolve, another member swirled the flask to get rid of any clinging powder. Next, we each slowly added the salt while swirling the flask. The solution was cloudy at first, because our team had the highest concentrate.

Results

The results after the first day were very different from what I predicted, Table 1 contains the number of crystals counted in three separate fields of view then averaged together, and my predictions. In the higher concentrates most of the crystals counted were educated guesses.

SAMPLE # 1 2 3 4 5 6 7 8
Prdicted 10 12 20 25 28 31 43 56
Actual 10 3 17 16 10 41 67 44

After looking at the observations, I suspected some type of bias because the numbers rose so steeply from sample five to sample six. And it dropped sharply from sample one sample two. The sizes of the crystals were very close to what I guessed. I got one size from three fields of view and averaged them and rounded them up.

SAMPLE # 1 2 3 4 5 6 7 8
Predicted .1mm .2mm .3mm .4mm .5mm .5mm .3mm .2mm
Actual .05mm .04mm .2mm .4mm .4mm .3mm .3mm .4mm

Most of the crystals were transparent and had light gray edges, and had these basic shapes:

drawing of crytals

The best sample so far is sample eight because it has the most and the largest protein crystals.

After the second day there were more and bigger crystals in the smaller concentrations. The largest crystals were in sample three and two. Samples five and six had the biggest increases in crystal production. But many of the other samples also had large increases.

SAMPLE # 1 2 3 4 5 6 7 8
Prdicted 12 18 25 24 15 50 75 56
Actual 19 22 46 12 62 90 50 63

There wasn't much of an increase in size, except in sample three and two. I used the same steps in averaging as the above examples.

SAMPLE # 1 2 3 4 5 6 7 8
Predicted .1mm .1mm .4mm .5mm .5mm .4mm .4mm .3mm
Actual .07mm .05mm .5mm .5mm .3mm .3mm .3mm .3mm

The largest crystals were in sample three. On day two most of the crystals were bulkier and more evenly distributed. They had just about the same color as on the first day, (transparent with light gray edges) but the crystal samples from three to eight were wider. The crystals included these shapes:

drawing of crytals

Discussion

Most of the sizes I predicted were very close to the actual sizes. But some samples still had one or two large .6mm to .7mm crystals like two occurrences in sample five that were.6mm and. 7mm. There were a few more occurrences in sample three and four too. The sizes went almost as planned the exceptions were samples one and two. The amount of crystals didn't go as well. Many of the samples surprised me they had incredible progress especially samples three, five and six. By the second day many of the samples were completely covered with crystals. The best protein sample so far is sample three because it has the largest crystals and large amount of them too. My hypothesis was proved incorrect, probably because the higher concentrations made more crystals but since they were all scrunched together they didn't grow as much. Meanwhile sample three had .35g of NaCl and it was able to produce a good amount of crystals but also gave them room to grow.

Career related to structural biology

A career I would like to have one day is a surgeon because I would be able to help people and I would know I made a difference. This career also sounds very interesting; it is something I won't get bored of easily. There is also a need of surgeons. Molecular biology can help me in this job because I would have a better understanding of how medicines and other therapeutic items are made.

If I were a surgeon I would make a difference in the world because I could cure people's wounds, and perform all types of surgeries depending on what type of doctor or surgeon I will be. Because accidents happen everywhere and most of the average Americans live a sedentary lifestyle a surgeon can really change someone by saving or bettering their lives.

This is isn't a very boring career either because it takes very hard work and concentration to go through medical school and learn new techniques and surgeries. It also takes a lot of guts and training to do the actual surgery, especially the first surgery. You also can't fall asleep in an appendectomy or heart surgery. Another tough lesson in becoming a surgeon is defeating the immense amount of pressure but the reward of helping someone is priceless.

Molecular biology can help in this aspect also because I'll get a better understanding in the way cells react to diseases. I could also know how they are made and their structural outline and components. I would need a basic understanding of proteins, cells and bacteria because these things can cause different reactions in our bodies.

There is also a need for surgeons because more and more people die and receive injuries from serious accidents everyday. If there were more doctors and surgeons there could be better treatment and more available doctors for the patients.

There are many important parts about being a surgeon but helping and saving lives is the most important. Even though this career is very challenging it is also very interesting. With the endless amounts of injuries there is a desperate call for more surgeons and with structural biology I can better understand the making of medicines and vital proteins and cells in the human body.