Quiz 1

•Grab a quiz sheet•Write down section #•Take everything off desk•You have 10 minutes…

1. What organelle functions in

energy production in animal

cells. What about plants cells?

2. Which of the following is

membrane bound?

Nucleus or Nucleoid

3. Which are the two modes of reproduction in Ciliates, such as Paramecium?

__________________________________

4. Trypanosoma belongs to the Phylum

_________________

5. This organism belongs to the Kingdom _________________ and moves by means of ____________

6. What is a unique feature of a parfocal microscope?

a) light is formed on the slideb) can use both eyes to look at an

imagec) focus is retained as you move

between objective lensd) one eye can be focused

independently of the other

7. How many structures of a protein are there?

SpectrophotometrySpectrophotometry

. Use of spectrophotometers.. Use of spectrophotometers.

. Zero the machine.. Zero the machine.

. Composition and use of a . Composition and use of a blank.blank.

. How to adjust wavelength.. How to adjust wavelength.

. Proper use of cuvettes.. Proper use of cuvettes.

ENZYME KINETICSENZYME KINETICS

ProteinsProteins

• A protein's function is due entirely to A protein's function is due entirely to its overall shape = its overall shape = conformation.conformation.

• Conformation is determined by 1˚, Conformation is determined by 1˚, 2˚, 3˚, and perhaps 4˚ structure of 2˚, 3˚, and perhaps 4˚ structure of the protein.the protein.

The primary The primary structure of a structure of a

proteinprotein

*sequence of *sequence of amino acidsamino acids

The The secondary secondary

structure of structure of a proteina protein

*Hydrogen *Hydrogen bonding bonding

between between C & NC & N

The tertiary The tertiary structure of structure of

a proteina protein

*Interactio*Interaction of amino n of amino acids and acids and

H2OH2O

The quaternary structure of proteinsThe quaternary structure of proteins*2 or more protein molecules joined together*2 or more protein molecules joined together

EEnzymesnzymes

• Enzymes are Enzymes are protein catalystsprotein catalysts..

• Lower the activation energy necessary Lower the activation energy necessary for a chemical reaction to occurfor a chemical reaction to occur..

• Act on a substrate, form an Act on a substrate, form an enzyme-enzyme-substrate complexsubstrate complex. Ultimately results in . Ultimately results in a product.a product.

• We are using barley amylase enzyme on We are using barley amylase enzyme on a starch substrate.a starch substrate.

Formation of an enzyme-substrate complexFormation of an enzyme-substrate complex

Environmental factors affecting enzyme activityEnvironmental factors affecting enzyme activity

Blanking spectrophotometerBlanking spectrophotometer-Set wavelength to Set wavelength to 560 nm560 nm

-With chamber empty, set Transmittance With chamber empty, set Transmittance T=0T=0 with with left knobleft knob

-Place blank cuvette in spectrophotometerPlace blank cuvette in spectrophotometer

-Set Absorbance Set Absorbance A=0A=0 with with right knobright knob

-Replace blank for experimental cuvetteReplace blank for experimental cuvette

-Take reading!!!Take reading!!!

BASIC PROCEDUREBASIC PROCEDURE

H O2

H O2

35 ml starch

35 ml

(or buffer)

5 ml

BLANK INITIAL

TIMED SAMPLES

1 ml enzyme

0.1 ml I KI2

1. Prepare 2 erlenmeyers (one for pH 1. Prepare 2 erlenmeyers (one for pH and one for temperature) and one for temperature)

2. Set wavelength to 560 nm2. Set wavelength to 560 nm

3. Prepare a blank and blank your spec3. Prepare a blank and blank your spec

4. Collect your initial reading (Time 4. Collect your initial reading (Time zero)zero)

5. TA will put 1 ml of enzyme in your 5. TA will put 1 ml of enzyme in your erlenmeyer.erlenmeyer.

6. Start timing…6. Start timing…

7. As the proper time interval 7. As the proper time interval approaches, swirl the flask to mix approaches, swirl the flask to mix and draw 5 ml of starch/enzyme up and draw 5 ml of starch/enzyme up into the pipette. You should into the pipette. You should pipette up the solution about 15 pipette up the solution about 15 sec before the reading time.sec before the reading time.

8. At the reading time, release the 8. At the reading time, release the solution into the cuvette and mix - solution into the cuvette and mix - the iodine stops the reaction the iodine stops the reaction immediately.immediately.

9. Readings should generally be 9. Readings should generally be taken within about 5 minutes.taken within about 5 minutes.

Experiments:Experiments:

• You will work in groups of three.You will work in groups of three.

• Each group will perform one run from the Each group will perform one run from the pHpH experiment (4.0, 4.5, 5.0, 5.5, 6.0 and 6.5)experiment (4.0, 4.5, 5.0, 5.5, 6.0 and 6.5)

• Then, each group will do one of the Then, each group will do one of the temperaturetemperature variables (15 variables (15º, 30º, 45º, 55º, º, 30º, 45º, 55º, 60º and 70º)60º and 70º)

pH experimentpH experiment

-Add 35 ml of buffer to 35 ml of starch -Add 35 ml of buffer to 35 ml of starch solution.solution.

-Blank is 1:1 dH-Blank is 1:1 dH22O:buffer (+ iodine)O:buffer (+ iodine)

Table 5-4. Absorbance readings over time of reactions at different pHs.

TIME (min)

pH 0 2 4 6 8 10 12 14 16 18 20

4.0

4.5

5.0

5.5

6.0

6.5

Temperature experimentTemperature experiment-Mix as in basic procedure. The reaction -Mix as in basic procedure. The reaction flask is kept in a water bath. flask is kept in a water bath.

-NOTE: Very cold and very hot solutions -NOTE: Very cold and very hot solutions might need to sit before being read.might need to sit before being read.

Table 5-3. Absorbance readings over time of reactions at different temperatures.

TIME (min)

Temp.(°C) 0 1 2 3 4 5 6 7 8 10 20

15

30

45

55

60

70

I will come around and add enzyme when your group

has…1. Finished basic procedure

1. 35 mL pH buffer + 35 mL starch2. Blank- 0.1 mL I + 2.5 mL pH + 2.5 mL DI H2O3. Placed 0.1 mL I in each cuvette

2. Blanked Spectrophotometer

3. Taken time = 01. Transfer 5 mL solution from flask into cuvette w/

I. Make sure to mix!

All of the data will be shared All of the data will be shared among the classamong the class

• You will record your data on the You will record your data on the computer. computer.

• A data set will be printed out for A data set will be printed out for each student.each student.

Reaction RateReaction Rate

= 1/2 change in absorbance / = 1/2 change in absorbance /

time for this change to occurtime for this change to occur

-There is a worksheet at the end of the chapter to help -There is a worksheet at the end of the chapter to help calculate reaction rate. calculate reaction rate.

- Additional copies of this worksheet can be printed off of Additional copies of this worksheet can be printed off of the homepage.the homepage.

- HHand this in with your lab report, so that I can see your and this in with your lab report, so that I can see your calculations.calculations.

- There is another handout ("How to Calculate Reaction There is another handout ("How to Calculate Reaction Rates") that is available from the lab homepage that Rates") that is available from the lab homepage that details the calculation of these rates.details the calculation of these rates.

2018161412108642000.0

0.5

1.0

1.5

Time (min)

Abs

orba

nce

i

f

i -

A -1/2 Ai

4 Figures

1. x-axis: Time *pHy-axis: Absorbance

2. x-axis: Time *Temp.y-axis: Absorbance

3. x-axis: pHy-axis: Rxn rate

4. x-axis: Temp.y-axis: Rxn rate

ENZYME KINETICSReaction Rate Calculation Sheet

* †

Temp.(°C) Ai Af ²A 1/2² A Ai-1/2²A TAi-1/2² A R.r. (*/†)

15

30

45

55

60

70

Lab report 1 (50 points)Lab report 1 (50 points)

• Refer to Appendix A - pp. 191-195 Refer to Appendix A - pp. 191-195 guidelines for writing a core lab report.guidelines for writing a core lab report.

• EcologyEcology has all journal articles prior to has all journal articles prior to 1996 on-line. 1996 on-line. http://www.jstor.org/journals/00129658.htmlhttp://www.jstor.org/journals/00129658.html

FiguresFigures

• These take some time to get right.These take some time to get right.

• Pay close attention to format.Pay close attention to format.

All lab reports will be

submitted to me AND

Turnitin.com ON TIME!!!

Attached to hard copy will

be 4 hand drawn figures

and signed academic

honesty form.

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Temperature ( C)

Chi

rp R

ate

(Hz)

Figure 1. Chirp frequency (Hz) of the striped ground cricket (Nemobius allardidifferent ambient air temperatures. The data presented are those of Pierce (1949).

) at

• Four graphs for this lab report:Four graphs for this lab report: 2 figures (Absorbance vs. time)2 figures (Absorbance vs. time)

• temperature vs. timetemperature vs. time• pH vs. timepH vs. time

2 figures (RR vs. T2 figures (RR vs. Tºº/pH)/pH)• Reaction rate vs temperatureReaction rate vs temperature• Reaction rate vs. pHReaction rate vs. pH

• Best fit curve for each set of dataBest fit curve for each set of data

2018161412108642000.0

0.5

1.0

1.5

Time (min)

Abs

orba

nce

For this first report, For this first report, ALL ALL FIGURES MUST BE DONE BY FIGURES MUST BE DONE BY HANDHAND (not by computer). In the (not by computer). In the future, you may use computer future, you may use computer generated figuresgenerated figures

(5) English• (3) Spelling, grammar, clarity• (2) Format

(1) Title(6) Abstract(12) Introduction

• (8) Background info• (4) Hypotheses

(6) Materials and Methods• (4) Experimental design and materials• (2) Data collection and analysis

(8) Results• (4) Tables and figures• (2) Stats• (2) Text

(10) Discussion• (8) Interpretation of data• (2) Alternative hypothesis/sources of error

(2) Literature cited• (1) All references cited in the text• (1) All references in text are cited

For next class

Quiz

• Bring charts to class or my office hours for me to look over!

• Read Ch 9 & 10 and “Animal Taxa” worksheet

• Bring dissection kits!