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Pamela El Khoury, Andy Awad, Brigitte Wex, Roy A. Khalaf
Proteomic analysis of a Candida albicans pir32 null strain reveals proteins involved in adhesion, filamentation and virulence
www.plosone.org
www.plosone.org
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3.3071 0 Td
(extraction)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 566.8157 cm
BT
/F9 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Twelve)Tj
3.0613 0 Td
(cell)Tj
1.5591 0 Td
(wall)Tj
1.8312 0 Td
(extractions)Tj
4.6204 0 Td
(from)Tj
2.2053 0 Td
(each)Tj
2.0296 0 Td
(strain)Tj
2.5002 0 Td
(and)Tj
1.7291 0 Td
(for)Tj
1.3833 0 Td
(growth)Tj
3.0727 0 Td
(condition)Tj
4.1159 0 Td
(were)Tj
2.126 0 Td
(performed)Tj
4.4674 0 Td
(inde-)Tj
-34.7015 -1.2982 Td
(pendently.)Tj
4.4333 0 Td
(Cells)Tj
2.16 0 Td
(were)Tj
2.1203 0 Td
(first)Tj
1.8085 0 Td
(centrifuged)Tj
4.7848 0 Td
(at)Tj
0.9525 0 Td
(4,000)Tj
2.347 0 Td
(rpm)Tj
1.9276 0 Td
(for)Tj
1.389 0 Td
(5)Tj
0.6916 0 Td
(min,)Tj
2.0806 0 Td
(then)Tj
2.0183 0 Td
(re-suspended)Tj
5.5842 0 Td
(in)Tj
1.0375 0 Td
(5)Tj
0.6973 0 Td
(mL)Tj
1.576 0 Td
(Tris)Tj
-35.6085 -1.3039 Td
(\(5)Tj
1.0318 0 Td
(mM,)Tj
2.1599 0 Td
(pH)Tj
1.5081 0 Td
(=)Tj
0.7993 0 Td
(7.8\).)Tj
1.9616 0 Td
(Protease)Tj
3.5943 0 Td
(Inhibitor)Tj
3.8551 0 Td
(Cocktail)Tj
3.5546 0 Td
(\(6)Tj
/F17 1 Tf
1.0375 0 Td
[()]TJ
/F9 1 Tf
(L,)Tj
1.491 0 Td
(abcam)Tj
2.8346 0 Td
(ab65621\))Tj
3.8835 0 Td
(along)Tj
2.4207 0 Td
(with)Tj
1.9956 0 Td
(cold)Tj
1.9219 0 Td
(glass)Tj
-34.0495 -1.2983 Td
(beads)Tj
2.4717 0 Td
(were)Tj
2.1203 0 Td
(added)Tj
2.6646 0 Td
(in)Tj
1.0318 0 Td
(a)Tj
0.6576 0 Td
(1:1)Tj
1.3947 0 Td
(beads)Tj
2.4718 0 Td
(to)Tj
1.0261 0 Td
(pellet)Tj
2.3754 0 Td
(volume)Tj
3.1975 0 Td
(ratio.)Tj
2.3244 0 Td
(Thirty)Tj
2.7496 0 Td
(cycles)Tj
2.5511 0 Td
(of)Tj
1.0148 0 Td
(vortexing)Tj
4.0252 0 Td
(were)Tj
2.1203 0 Td
(applied)Tj
-34.1969 -1.2983 Td
(to)Tj
1.0261 0 Td
(ensure)Tj
2.8743 0 Td
(breakage)Tj
3.7757 0 Td
(as)Tj
1.0148 0 Td
(follows:)Tj
3.2825 0 Td
(30)Tj
1.1736 0 Td
(sec)Tj
1.4286 0 Td
(on)Tj
1.2756 0 Td
(vortex)Tj
2.7382 0 Td
(followed)Tj
3.651 0 Td
(by)Tj
1.1792 0 Td
(30)Tj
1.1736 0 Td
(sec)Tj
1.423 0 Td
(on)Tj
1.2755 0 Td
(ice.)Tj
1.5648 0 Td
(Samples)Tj
3.4922 0 Td
(turned)Tj
-32.3487 -1.3039 Td
(orange)Tj
2.9706 0 Td
(reflecting)Tj
3.9742 0 Td
(a)Tj
0.652 0 Td
(reaction)Tj
3.4922 0 Td
(between)Tj
3.532 0 Td
(the)Tj
1.4683 0 Td
(acidic)Tj
2.5625 0 Td
(cytosol)Tj
3.016 0 Td
(and)Tj
1.7292 0 Td
(the)Tj
1.474 0 Td
(Protease)Tj
3.5943 0 Td
(inhibitor.)Tj
4.0081 0 Td
(Beads)Tj
2.5455 0 Td
(were)Tj
-35.0189 -1.2982 Td
(then)Tj
2.0069 0 Td
(removed)Tj
3.7417 0 Td
(by)Tj
1.1678 0 Td
(pouring)Tj
3.4186 0 Td
(the)Tj
1.4627 0 Td
(samples)Tj
3.3732 0 Td
(into)Tj
1.8311 0 Td
(other)Tj
2.3358 0 Td
(pre-weighed)Tj
5.193 0 Td
(tubes)Tj
2.3301 0 Td
(and)Tj
1.7234 0 Td
(washing)Tj
3.498 0 Td
(the)Tj
1.457 0 Td
(beads)Tj
2.4661 0 Td
(sev-)Tj
-36.0054 -1.2983 Td
(eral)Tj
1.6837 0 Td
(times)Tj
2.3811 0 Td
(with)Tj
1.9899 0 Td
(cold)Tj
1.9162 0 Td
(NaCl)Tj
2.2904 0 Td
(until)Tj
2.0976 0 Td
(the)Tj
1.4627 0 Td
(supernatant)Tj
4.9719 0 Td
(becomes)Tj
3.6794 0 Td
(transparent;)Tj
5.0399 0 Td
(making)Tj
3.2429 0 Td
(sure)Tj
1.8935 0 Td
(not)Tj
1.5647 0 Td
(to)Tj
1.0148 0 Td
(trans-)Tj
-35.2287 -1.3039 Td
(fer)Tj
1.3039 0 Td
(any)Tj
1.6554 0 Td
(beads)Tj
2.4718 0 Td
(to)Tj
1.0261 0 Td
(the)Tj
1.4684 0 Td
(new)Tj
1.8765 0 Td
(pre-weighed)Tj
5.2044 0 Td
(tubes)Tj
2.3357 0 Td
(during)Tj
2.9253 0 Td
(the)Tj
1.474 0 Td
(process.)Tj
3.4073 0 Td
(The)Tj
1.7801 0 Td
(efficiency)Tj
4.0309 0 Td
(of)Tj
1.0148 0 Td
(breakage)Tj
3.7757 0 Td
(was)Tj
-35.7503 -1.2983 Td
(examined)Tj
4.1272 0 Td
(under)Tj
2.6192 0 Td
(the)Tj
1.4683 0 Td
(microscope.)Tj
5.0683 0 Td
(The)Tj
1.7802 0 Td
(samples)Tj
3.3845 0 Td
(were)Tj
2.1203 0 Td
(spun)Tj
2.177 0 Td
(at)Tj
0.9524 0 Td
(3,000)Tj
2.3471 0 Td
(rpm)Tj
1.9276 0 Td
(for)Tj
1.3889 0 Td
(5)Tj
0.6917 0 Td
(min,)Tj
/F16 1 Tf
2.0806 0 Td
(t)Tj
/F9 1 Tf
(he)Tj
1.5137 0 Td
(superna-)Tj
-33.647 -1.2983 Td
(tants)Tj
2.1599 0 Td
(containing)Tj
4.5241 0 Td
(intracellular)Tj
5.0626 0 Td
(proteins)Tj
3.515 0 Td
(were)Tj
2.1203 0 Td
(poured)Tj
3.0954 0 Td
(off,)Tj
1.525 0 Td
(while)Tj
2.3698 0 Td
(pellets)Tj
2.7382 0 Td
(were)Tj
2.1203 0 Td
(re-suspended)Tj
5.5842 0 Td
(in)Tj
-34.8148 -1.2982 Td
(NaCl)Tj
2.296 0 Td
(\(40)Tj
1.5137 0 Td
(mL,)Tj
1.8028 0 Td
(1)Tj
0.6973 0 Td
(M\))Tj
1.4514 0 Td
(and)Tj
1.7291 0 Td
(spun)Tj
2.177 0 Td
(again)Tj
2.3697 0 Td
(at)Tj
0.9525 0 Td
(3,000)Tj
2.347 0 Td
(rpm)Tj
1.9276 0 Td
(for)Tj
1.3833 0 Td
(5)Tj
0.6973 0 Td
(min.)Tj
2.0806 0 Td
(This)Tj
1.9899 0 Td
(NaCl)Tj
2.2961 0 Td
(washing)Tj
3.5092 0 Td
(step)Tj
1.8199 0 Td
(was)Tj
-33.0404 -1.3039 Td
(repeated)Tj
3.6396 0 Td
(three)Tj
2.262 0 Td
(to)Tj
1.0262 0 Td
(four)Tj
1.9162 0 Td
(times.)Tj
2.6135 0 Td
(Protein)Tj
3.1918 0 Td
(extraction)Tj
4.2576 0 Td
(buffer)Tj
2.6249 0 Td
(\(50)Tj
1.5137 0 Td
(mM)Tj
1.9332 0 Td
(Tris,)Tj
2.0523 0 Td
(2%)Tj
1.457 0 Td
(SDS,)Tj
2.1316 0 Td
(100)Tj
1.6441 0 Td
(mM)Tj
1.9332 0 Td
(Na-)Tj
-34.1969 -1.2983 Td
(EDTA,)Tj
3.0443 0 Td
(150)Tj
1.6498 0 Td
(mM)Tj
1.9275 0 Td
(NaCl,)Tj
2.5285 0 Td
(pH)Tj
1.5024 0 Td
(7.8\))Tj
1.7404 0 Td
(with)Tj
/F15 1 Tf
1.9956 0 Td
()Tj
/F9 1 Tf
(-mercaptoethanol)Tj
7.8349 0 Td
(\(8)Tj
/F17 1 Tf
1.0375 0 Td
[()]TJ
/F9 1 Tf
(L)Tj
1.2642 0 Td
(per)Tj
1.5307 0 Td
(1)Tj
0.6974 0 Td
(mL)Tj
1.576 0 Td
(protein)Tj
3.1578 0 Td
(extraction)Tj
-31.487 -1.2983 Td
(buffer\))Tj
2.965 0 Td
(was)Tj
1.7007 0 Td
(added)Tj
2.6589 0 Td
(\(0.5)Tj
1.7348 0 Td
(mL)Tj
1.5817 0 Td
(buffer)Tj
2.6249 0 Td
(per)Tj
1.5307 0 Td
(100)Tj
1.6441 0 Td
(mg)Tj
1.508 0 Td
(wet)Tj
1.6271 0 Td
(weight)Tj
2.8856 0 Td
(walls\))Tj
2.5285 0 Td
(and)Tj
1.7292 0 Td
(the)Tj
1.474 0 Td
(pellets)Tj
2.7325 0 Td
(were)Tj
2.126 0 Td
(re-sus-)Tj
-33.0517 -1.3039 Td
(pended.)Tj
3.4128 0 Td
(Tubes)Tj
2.6476 0 Td
(were)Tj
2.126 0 Td
(boiled)Tj
2.6985 0 Td
(for)Tj
1.3833 0 Td
(10)Tj
1.1736 0 Td
(min)Tj
1.8538 0 Td
(and)Tj
1.7291 0 Td
(spun)Tj
2.177 0 Td
(for)Tj
1.3833 0 Td
(5)Tj
0.6973 0 Td
(min)Tj
1.8539 0 Td
(at)Tj
0.9524 0 Td
(3,000)Tj
2.3528 0 Td
(rpm.)Tj
2.1486 0 Td
(The)Tj
1.7858 0 Td
(supernatants)Tj
5.3405 0 Td
(con-)Tj
-35.7163 -1.2982 Td
(taining)Tj
3.05 0 Td
(the)Tj
1.4683 0 Td
(SDS)Tj
1.9049 0 Td
(extractable)Tj
4.5468 0 Td
(proteins)Tj
3.5149 0 Td
(were)Tj
2.1203 0 Td
(collected)Tj
3.7417 0 Td
(and)Tj
1.7291 0 Td
(later)Tj
1.9956 0 Td
(subjected)Tj
3.9685 0 Td
(to)Tj
1.0205 0 Td
(tryptic)Tj
2.8516 0 Td
(digestion)Tj
3.8891 0 Td
(and)Tj
-35.8013 -1.2983 Td
(subsequent)Tj
4.7054 0 Td
(steps)Tj
2.177 0 Td
(for)Tj
1.389 0 Td
(protein)Tj
3.1521 0 Td
(identification.)Tj
5.811 0 Td
(Protein)Tj
3.1974 0 Td
(extraction)Tj
4.2577 0 Td
(buffer)Tj
2.6248 0 Td
(and)Tj
/F15 1 Tf
1.7291 0 Td
()Tj
/F9 1 Tf
(-mercaptoethanol)Tj
-29.0435 -1.3039 Td
(were)Tj
2.1203 0 Td
(added)Tj
2.6588 0 Td
(again)Tj
2.3698 0 Td
(as)Tj
1.0148 0 Td
(before)Tj
2.7439 0 Td
(to)Tj
1.0205 0 Td
(re-suspend)Tj
4.6374 0 Td
(pellet.)Tj
2.5965 0 Td
(Samples)Tj
3.498 0 Td
(were)Tj
2.1259 0 Td
(boiled,)Tj
2.9197 0 Td
(cooled,)Tj
3.0841 0 Td
(centrifuged)Tj
4.7792 0 Td
(for)Tj
1.3889 0 Td
(5)Tj
-36.9578 -1.2983 Td
(min)Tj
1.8538 0 Td
(at)Tj
0.9524 0 Td
(3,000)Tj
2.3471 0 Td
(rpm,)Tj
2.1543 0 Td
(and)Tj
1.7291 0 Td
(re-suspended)Tj
5.5843 0 Td
(in)Tj
1.0374 0 Td
(water.)Tj
2.6532 0 Td
(Wash)Tj
2.5115 0 Td
(steps)Tj
2.1827 0 Td
(with)Tj
1.9956 0 Td
(Type)Tj
2.228 0 Td
(1)Tj
0.6916 0 Td
(water)Tj
2.4321 0 Td
(were)Tj
2.126 0 Td
(done)Tj
2.2224 0 Td
(to)Tj
-34.7015 -1.2983 Td
(remove)Tj
3.2201 0 Td
(excess)Tj
2.6759 0 Td
(SDS.)Tj
2.1316 0 Td
(The)Tj
1.7858 0 Td
(final)Tj
2.0013 0 Td
(pellets)Tj
2.7382 0 Td
(were)Tj
2.1203 0 Td
(frozen)Tj
2.7723 0 Td
(in)Tj
1.0375 0 Td
(liquid)Tj
2.5625 0 Td
(N)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 437.8393 305.1212 cm
BT
/F9 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(2)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 443.5653 306.822 cm
BT
/F9 1 Tf
1 TL
-0.004 Tc
0 0 Td
(and)Tj
1.7291 0 Td
(freeze-dried.)Tj
5.2554 0 Td
(Lyophilized)Tj
4.9095 0 Td
(cell)Tj
-36.2491 -1.3039 Td
(walls)Tj
2.1884 0 Td
(were)Tj
2.1259 0 Td
(finally)Tj
2.7043 0 Td
(stored)Tj
2.7099 0 Td
(at)Tj
0.9524 0 Td
(-20C)Tj
2.5738 0 Td
(until)Tj
2.109 0 Td
(use.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 266.5133 cm
BT
/F16 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(2.4)Tj
1.4598 0 Td
(Extraction)Tj
4.6063 0 Td
(of)Tj
1.0678 0 Td
(alkali)Tj
2.4992 0 Td
(labile)Tj
2.4898 0 Td
(cell)Tj
1.6346 0 Td
(wall)Tj
1.9229 0 Td
(proteins)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 249.5622 cm
BT
/F9 1 Tf
1 TL
-0.004 Tc
0 0 Td
(The)Tj
1.7801 0 Td
(cell)Tj
1.559 0 Td
(wall)Tj
1.8312 0 Td
(pellets)Tj
2.7383 0 Td
(were)Tj
2.1203 0 Td
(subjected)Tj
3.9684 0 Td
(to)Tj
1.0262 0 Td
(overnight)Tj
4.0818 0 Td
(incubation)Tj
4.5525 0 Td
(with)Tj
1.9955 0 Td
(NaOH)Tj
2.9027 0 Td
(\(30)Tj
1.5137 0 Td
(mM\))Tj
2.2734 0 Td
(at)Tj
0.9524 0 Td
(4C.)Tj
1.9729 0 Td
(They)Tj
-35.2684 -1.3039 Td
(were)Tj
2.1203 0 Td
(then)Tj
2.0182 0 Td
(neutralized)Tj
4.6998 0 Td
(with)Tj
2.0013 0 Td
(aqueous)Tj
3.5036 0 Td
(acetic)Tj
2.4944 0 Td
(acid)Tj
1.8709 0 Td
(\(30)Tj
1.508 0 Td
(mM\))Tj
2.2734 0 Td
([)Tj
0.83 0.64 0.02 0 k
(18)Tj
0 g
(].)Tj
2.0806 0 Td
(Samples)Tj
3.4923 0 Td
(were)Tj
2.1259 0 Td
(spun)Tj
2.1714 0 Td
(at)Tj
0.9524 0 Td
(3,000)Tj
2.3527 0 Td
(rpm)Tj
-35.6652 -1.2982 Td
(for)Tj
1.3832 0 Td
(5)Tj
0.6974 0 Td
(min,)Tj
2.0806 0 Td
(and)Tj
1.7291 0 Td
(supernatants)Tj
5.3461 0 Td
(were)Tj
2.1203 0 Td
(collected)Tj
3.7417 0 Td
(and)Tj
1.7291 0 Td
(subjected)Tj
3.9685 0 Td
(to)Tj
1.0262 0 Td
(tryptic)Tj
2.8516 0 Td
(digestion.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 196.2708 cm
BT
/F16 1 Tf
1 TL
-0.0033 Tc
0 0 Td
(2.5)Tj
1.4598 0 Td
(Glucanase)Tj
4.4977 0 Td
(treatment)Tj
4.3748 0 Td
(of)Tj
1.063 0 Td
(cell)Tj
1.6347 0 Td
(wall)Tj
1.9228 0 Td
(pellets)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 200.0125 179.2629 cm
BT
/F9 1 Tf
1 TL
-0.004 Tc
0 0 Td
(Spectrophotometric)Tj
8.1977 0 Td
(analysis)Tj
3.3222 0 Td
(was)Tj
1.7008 0 Td
(used)Tj
2.0579 0 Td
(to)Tj
1.0261 0 Td
(estimate)Tj
3.549 0 Td
(cell)Tj
1.559 0 Td
(numbers.)Tj
4.0025 0 Td
(10)Tj
ET
Q
q
1 j
1 J
0 w
7.4999 0 0 6.6659 463.6913 183.7417 cm
BT
/F9 1 Tf
1 TL
-0.0053 Tc
0 0 Td
(8)Tj
ET
Q
q
1 j
1 J
0 w
10 0 0 10 469.4173 179.2629 cm
BT
/F9 1 Tf
1 TL
-0.004 Tc
0 0 Td
(cells)Tj
1.9218 0 Td
(were)Tj
2.126 0 Td
(incubated)Tj
4.1726 0 Td
(at)Tj
-35.1607 -1.2982 Td
(37C)Tj
2.2281 0 Td
(with)Tj
1.9955 0 Td
(1)Tj
0.6974 0 Td
(mg)Tj
1.5023 0 Td
(of)Tj
1.0148 0 Td
(Glucanase)Tj
4.3313 0 Td
(\()Tj
/F15 1 Tf
()Tj
/F9 1 Tf
(-\(13\)-D-Glucana)Tj
8.0957 0 Td
(se)Tj
1.0035 0 Td
(from)Tj
/F8 1 Tf
2.2053 0 Td
[()31()15()12()]TJ
2.3187 0 Td
[(\n)4()11()6()22()7()4()]TJ
/F9 1 Tf
(,)Tj
3.702 0 Td
(Sigma-Aldrich\))Tj
6.4176 0 Td
(in)Tj
-35.5122 -1.2983 Td
(sodium)Tj
3.2315 0 Td
(acetate)Tj
2.9537 0 Td
(buffer)Tj
2.6249 0 Td
(\(1)Tj
1.0374 0 Td
(mL,)Tj
1.8029 0 Td
(150)Tj
1.6441 0 Td
(mM,)Tj
2.1599 0 Td
(pH)Tj
1.5024 0 Td
(5\))Tj
1.0375 0 Td
(overnight)Tj
4.0818 0 Td
([)Tj
0.83 0.64 0.02 0 k
(19)Tj
0 g
(].)Tj
2.075 0 Td
(Supernatants)Tj
5.4594 0 Td
(were)Tj
2.1259 0 Td
(collected)Tj
3.7361 0 Td
(and)Tj
-35.4725 -1.3039 Td
(subjected)Tj
3.9685 0 Td
(to)Tj
1.0262 0 Td
(tryptic)Tj
2.8516 0 Td
(digestion.)Tj
ET
Q
q
1 j
1 J
0 w
11.9999 0 0 11.9999 200.0125 112.9889 cm
BT
/F16 1 Tf
1 TL
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