Gram Stain and Peptidoglycan Flashcards
first step of gram stain
primary stain (crystal violet) is added to heat-fixed/dead bacteria
second step of gram stain
add grams iodine (acts as a mordant) - binds crystal violet, making it a large molecule so that it is very difficult for it to leave the ppg layer
third step of gram stain
add decolorizer (wash step), which is acetone or ethanol. this will wash out all of the crystal violet except that which is in the ppg layer and cannot leave due to the binding of the iodine
fourth step of gram stain
secondary stain (safranin/red), which turns anything that is colorless pink
describe results of gram stain
gram + cells will be purple because the crystal violet will highlight the thick ppg layer. gram - cells will be red because the crystal violet does not get stuck in the ppg and therefore washes away, leaving the safranin to stain them pink in the last step
components and structure of peptidoglycan (ppg)
- carbohydrates: NAG and NAM
- tetrapeptides: chains of 4 amino acids
- backbone of the molecule is repeating NAG and NAM with tetrapeptides coming off of each NAM
- in gram - bacteria, tetrapeptides are cross-linked by covalent bonds
- in gram + bacteria, tetrapeptides are linked by additional amino acids, making the ppg thicker in these cells
- ppg is helical
what does penicillin do
penicillin interferes with ppg synthesis by preventing cross-linking of adjacent ppg chains. it is usually more effective against gram + bacteria. specifically, penicillin has B-lactam rings that bind to enzymes that crosslink the tetrapeptides
what does a lysozyme do
it is an enzyme found in tears and saliva, breaks NAG/NAM glycosidic bonds
what is different about the mycoplasma species?
they do not have a cell wall/ppg and have extremely variable shapes (pleomorphic). their membrane contains sterols that increase strength and they are not affected by penicillins
describe archaea cell walls
pseudo ppg, cell walls extremely variable due to extreme environments in which they live
