Indian Journal of Biochemistry & Biophysics Vol. 38. June 200 I, pp. 153- 158

A model for the organization of chlorophyll-protein complex of photosystem II and analysis of its photochemical efficiency and excitation migration

L S Ralh and M K Raval*

Post GraduJte DepJrlment of Chcmi stry. RajcndrJ Co ll ege, Bolangir 767002, OrissJ, India

Recei\'ed 14 Seplelllbcr 2000; rel'ised alld accepled 6 Decelllber 2000

A Illude I is proposed for the orga ll izat ion of chlorophyll-protei ll complex in photosystem II (PS II ) of highcr plants. The ra tes 01" excitoll mi grJtion and exc iton trapp ing have been cO lllputed using stochasti c method to find out the photochemical el"ficiency of the dimeric PS 11. Th ree dimeric PS II un its are assumed to form a group, JS trJnsf(:'r of the exc iton 10 the ligh t harvesti ng bed of the nearest neighbour on eit her side may on ly be effec tive. A relationshi p has been deduced hetween the I"ract ions of the reJcti on ce nt re traps closed and the number of jumps (J) req uired by the exc iton for trapping. The photochem ica l erl"iciency and fl uorescence qU Jn lllm yield are computed using J as the parameter in an empirical equation.

Photosystel1l II (PS II) is a protein complex of thylakoid that utilizes the light energy to split water to molecular oxygen. It is located across the lipid bilayers of the thylakoid membrane of the chloroplast. It consists of three major components I ·) : (1) the reaction centre II (RC II), (2) the antenna proteins and (3) the light harvesting chlorophyll alb binding proteins (LHC II) . The RCII consists of the D I and D2 proteins-l·C,. The D I/D2 hetrodimer provides a matrix for a special pair of chlorophyll a (ChI a)l molecules. pheophytins and electron acceptor (p lastoquinones QA and QD) responsible for the electron transport across the thylakoid membrane. The 43 kDa and 47 kDa antenn a protei ns contai ning 12 ChI a and 5 ~-caro te nes each, are located surrounding the RC II (ref. 7). These antenna protei ns channelize the light energy from the outermost LHC II proteins to the RC II. Carotenoids have a very low effici ency in energy transfer to RC II and thei r major ro le is protection of the system from photodamage through the quenching of singlet oxygen8•9 • The LHC II proteins are present surrounding the antenna proteins. The function of these proteins is to harvest the light energy and transfer it to the antenna proteins which in turn transfer it to the RCII. The spec ial pair of (ChI ah acts as a photon trap, i.e., photon absorbed by the (ChI ah is not migrated back to other pigments. The trap converts photon to photochemi cal energy by the primary charge separation lead ing to the formation of P680+. In thi s state the trap is known to be "closed"

* Author for correspondence

a~ it can not absorb another exciton. When it is reduced back to form P680 it is ready to absorb an exciton, hence in th is state the trap is designated as "open". Earl ier we have proposed a model of monomeric PS II for exciton migration based on the stoichiometry of pigments and polypeptides lU. However, at present the info rmation on the structural organization of PS II has increased . The knowledge of a three dimens ional (3D) structure of PS II at higher resolution can give a deeper insight into the phenomena associated with energy transfer. Recently two and three-di mensional structures of plant PS l1 at low resolutions have been determ i n ed' ·J· II .,~. An electron crystallograph ic structure of a LHC II protein , Lhcb I (fllcb ) gene prod uct) has been determined at 3.4 A resol uti on revea ling the organizat ion of Chi alb in it. However, the detailed st ructure of the antenna proteins and the RC II at hi gher reso lution is awaited which can project a clear picture of the proteins at atomic level. Changes in the organi zation and size of LHC 11 around antenna proteins are believed to be reflec ted by the fl uorescence quantum yield and the rate of O~ evolution in PS n. However, a quant itati ve relationshi p between the size and organi zati on of LHC II and the quantum yield is not avai lable. Such a relationship would be useful to predict the adaptational changes of LHC II organi zation under vari ous light and stress conditions by look ing at the quantum yield parameters.

In the present paper, we have made an attempt to modify our previous model on the basis of the recent

154 INDIAN J. BIOCHEM . 1310PHYS., VOL. 38, JUNE 200 1

experimental repOIts on the structure of PS I1 1-3.IO. IJ- 17. The photochemical efficiency, excitation transfer and Iluorescence qu antum yield are computed by lIsing stati stical methods. The result agrees well with the

. I f I 18 19 I . ex pen menta reports 0 t lese parameters . . t IS an in iti al step in search for a quantitati ve relationship

Fig. 1- Schematic diagram of the proposed model of dimeric PSII showing organi zati oI1J of various proteins. [LH2 represents Lhcb I, Lhcb2 or Lhcb3 proteins; LH I represents Lhcb4, Lhcb5 or Lhcb6 protein s. Antenna proteins are designated as 43 kDa and 47 kDa. D I and D2 are reaction centre II proteins. Modeled after Rhee e l al. 2 and Eijckelhoff el a16 J.

between the st ructure and quantum yield parameters. The rate of exci ton migration i.e., the number of jumps (1) an exciton makes to reach the trap with probability 0.99 depends on the structural organi zation of the LHC II , the an tenna and the trap. The rate of exci ton migration determines the va lues of fluorescence quantum yield or the rate of O2 evolution. An equation is derived in the present work to establi sh a quantitati ve relationship between J and quantum yield parameters.

The Model The proposed model of LHC II is based on the

reported stoichiometry and knowledge of organ ization of proteins in PS II )·3.6.9.20 (Fig. I ). The structure of each LHC II polypeptide is homologous to Lhcb I (ref. 21). The two dimensional electron crystallo-graph ic structure of pea Lhcb I at 3.4 A reveal s the presence of 12 Chi alb molecules per LHC II. [n the present model (Fig. I ) we have taken 12 Chi alb molecules per LHC II . It is proposed th at 12 LHC " are attached on each side of the dimer of D I/D2, Cyt b559, 43 kDa and 47 kDa (Fig. I ). The total number of Chi alb associated with each PS II is [44 (Fig.2). This makes the outermost layer i.e. , layer 1 of Chis. The antenn a proteins -43 kDa and 47 kDa are present on the inner side of LHC II. Each antenna protein is predicted to bind to 12 Chi a molecules 7• Thus 24 Chi a per PS II are present in the second layer of the organizati on. The RC II contains a special pair of (Chi ah i. e., P680, which acts as the exc iton trap. It makes the third layer (Fig.2). Thus the dimeric PS II model contains an array of 288 Chi alb molecules in layer I, 48 Chi a molecules in layer 2 and the P680 of trap 1 and trap 2, making layer 3 and layer 4 respectively, each containing two Chi a molecules.

Fi g. 2-Schemati c diagram showing a domain of three dimeric PS II units. [The domain consists of ten sets of chlorophy ll pi gments. NUlll ber of chlorophyll molecules in each set is indicated] .

RATH & RAVA L: A MODEL FOR CHLOROPHYLL-PROTEIN COMPLEX OF PHOTOSYSTEM-II 155

The exciton transfer between the pigments of a polypeptide unit is assumed to be coherent as the pigments are at a di stance of about loA from one another and at such a small di stance the molecular orbitals can interact with each other leading to strong couplingl l. The migration of exciton from one polypeptide unit to the nearest neighbours is assumed to be Foster-resonance type22-25 . The model for energy transfer is a si mple version of the experimentall y observed complex phenomenon under ill vivo condi tion26. In the LHC II energy transfers between Chi b ---7 Chi Q are observed in multiphasic time scale, e.g., 180 fs, 600 fs and 2-5 pS27. Li ght induced act ivation of LHC II is suggested to occur via a photoconductive mechani sm28 . The li ght induced enhancement of electron density in LHC II indicates an increase in energy and delocalizati on of electron density which is important for effici ency and speed of energy transfer from LHC II to the RC II. The time scale of 10-20 ps fo r intermonomer energy transfer supports the Foster type transfer proposed in the present model29. From layer I , exciton can jump to the neighbouring layer 2 only and not to any other layer. Similarly fro m layer 2, exciton can either jump to the neighbouring layer I or layer 3 (or layer 4). The layer 3 and layer 4 are the exciton traps where the light energy is converted to the photochemical energy. It is assumed that the exciton does not escape the trap unless it is closed.

In our theoretical model, three dimeric PS II centres are clubbed to form a group with a common LHC bed. The distribution thus gives ten sets of pigments (Fig.2). Clubbing of three PS II units are based on an approx imation that for an effective trapping an exciton may jump to its nearest neighbour on either side in a linear array of the layers. Such groupings are supported by ex perimental results I9. 30.

Methods An excitation transfer probability matri x, M, is

constructed. For ten sets of pigments, M would be a lO x 10 squ are matrix. The element, Mij , of the matrix M, when i = j , represents the probability that photon absorbed by set i will be retained by the same set of pigments. When i -:f:. j, Mij , represents the relative probability of exciton tranfer from set i to set j. The clements Mij of M are determ ined by counting the total number of possible transfers from ith set and the r . t·· I I d f' ·th 72?5 'rl ractlon 0 It t 1at ca · s to trans er to J set'· -. le rate of transfer from one set to the other is also proportional to a factor k 2 whi ch is 1 for parallel

orientation of the dipole moments of the Chi Q and 2/3 for random orientations22 . In the present model, k" is taken as 2/3 assuming an overall random orientat ion of ChI molecules . Again number of the transfer from one set to the other is multiplied by a weightage factor and normalized considering the different absorption maxima of pigments in differen t sets22 . Probability of transfer will be more from a set absorbi ng at lower wavelength to the one absorbing at a higher wave length. In the present model LHC II , antenna and trap fo rm the three different sets of pigments with three different absorption max ima 667, 675 and 680 nm respectively22. The weightage factors are adapted from Seely (1973)22. The factors are 0.1734 for exciton migration from a set to an outside set with lower absorption wavelength, 0.3564 for residing in the same set and 0.4702 for migrat ion to the inner set with higher absorption wavelength . Once the exciton hits an open trap, the probability of retention of the exci ton is 1.0 and the exci ton is said to be trapped. The probability of exciton di stribution is given by a lO x I column matrix , P. The initial values of elements of P are the ratios of the number of Ch i molecules in each set to the total number of pigments . The sum of elements of P is adjusted to 1.0000.

The probability of exciton distribution after J jumps is given by MjP. The value of J is recorded, when the probability of trapping of exciton reaches 0.99. The trapping rate is expressed in terms of the number of jumps, J, required to atta in a probability of trapping equal to 0.99.

Results and Discussion Seven diffe rent exci ton transfer probability

matrices, M(O), M(l), M(2), M(3), M(4) , M(5) and M(6) are constructed with 0, I , 2, 3, 4, 5 or 6 traps closed, respectively, in the domain (Fig. 2) . When all the six traps are open, the number of jumps (Jo) for trapping is found to be 62. When all the traps arc closed then J tends to infinity. The result is given in Table I.

Fraction of traps closed (x) has a pos iti ve correlation with In J (r = 0.99). Therefore, the fo llowing logarithmic eq uation for jump (J ) and fraction of traps closed (x) is proposed whi ch yi elds a constant value at x = 0 and infinity at x = I.

J=[A(exp(ax»]/( I-x)b . .. (I )

Where A is the constant giving the number of jumps required for trapping when all the six traps are open and a, b are constants.

156 INDIAN J. B10CI-IEM. 13I0PI-IYS .• VOL. 38, JUN E 2001

Table 1- Number of jumps req uired for an exciton before being trapped with probabi lity. p. as computed by stochastic method

Number of traps open

6 5 4 3 2 I o

Number of traps closed

o I 2 3 4 5 6

]t can be written in a linear form as

In J = In A + ax - b In (I - x)

Fraction of traps closed

(x)

0.00 0.16 0.33 0.50 0.66 0.84 1.00

... (2) The multiple regression gives the values of the

constants a, b and A (R2 = 0.9955, standard enor = 0.1179). The values are, a= 1.0156±0.2561 (t=3.966), b=0.6979 ±0.0575 (t= - 12.13), In A= 4.1679±0.0881 (1=47.229). The values calculated from the equation and calcul ated by matrix agree well (Table I, Fig.3). Number of jumps required for an exci ton to be trapped is calculated for various x values using the equation (1).

At x = I , theoretically J tends to infinity, but practically the number of jumps will be finite. Because the energy will not be used in the oxygen evolution when all the traps are closed and the exciton can keep on jumping for its life time which is approx imately 4.5 ns, 'r (0), under that circumstance31. If each jump takes one ps, 'r (1) then the maxi mum numbers of jumps (1maX> will be restricted to

Jmax = 'r (0) / 'r (1) = 4.5 X 10-9 sll X 10-12 s = 4500 . .. (3)

The fluorescence quantum yield (

RATH & RA V AL: A MODEL FOR CHLOROPHYLL-PROTEIN COMPLEX OF PHOTOSYSTEM-II 157

The quantitative relationships in our previous model was empirically proposed 10, however, in this paper we have improved them by deriving the equations through statistical methods . The monomeric PS )[ in our previous model shows lower quantum efficiency as compared to the dimeric one in the present model. The value of Jo i.e., the number of jumps required to reach the trap with a probability of 0.99, when all traps are opened for the previous model was 128, whereas in the present model it is 65. This suggests that the dimeric PS II organization might have been adapted by nature to achieve a better quantum effici ency.

The calculated values of quantum yield parameters in the present model are in good agreement with the experimental values shown in Fig. 4 (X2 for il¢rel is

0.4593 and X2 for V(02)rel is 0.0222). The number of degrees of freedom is 10. The calculated X2 values are extremely smaller than any of the tabulated X2 values. Therefore, V (02)rcl and il¢rel values calculated from the model are not significantly different from experimental values. Thi s method can be used to

calculate il¢rel and V(02)rel for changes in organization of LHCs in PS rl. The reduction in size of LHC [[ will influence matrix P, whereas uncoupling of LHC II and 43, 47 kDa proteins will affect the matrix M and hence the rate of transfer of

~ 1.0~~"~ .• ~.-.------------------------------~

ON

C ~ 0.8 ';;' c: Q)

OJ >-~ 0.6 --~ ., e~

.2 ~ 0.4 c: Q)

o til

~ o ~ 0.2 Q)

£ !:! Q)

•• '. •• •

o • o

o

~ o~----~------~------__ ----~------~ o 0.2 0.4 0.6 0.8 1. 0

Fraction of traps closed

Fig. 4 - Graph show ing the reiJtive rate of molec ul ar oxygen evolution. V(O~)rcl ( . --. ) and relati ve nuorescence qua ntum yield , 6rcl ( D--O ) I 'USIIS the fracti on of traps closed (x), using the predi cted mathematical formul a (see tex t for equat ions 7 and 8). It is compared with experimentall y determined V(02),cl ( ..... .. .... . ~. ) and 6

158 INDIAN J. BlOCl-l EM. BIOPHYS ., VOL. 38, JUNE 2001

26 Fleming R G & van Grondelle R (1997) Cllrr Opill Struct Bioi 7,738-748

27 Connaelly J P. Muller M G, Hucks M, Gatzen G, Mu llineau C W, Ruban A V, Herton P & Holzwarth A R (1997) J Phys G elll BIOI, 1902-1909

28 Lu kins P B ( 1999) Biochelll Biophys Res COli 1111 1111 256(2), 288-292

29 Visser H M, Kleima F J, van Stokkum I H M, van Grondell e R & van Amerongen H (1996) Chelll Phys 2 10, 297-312

30 Srivastava A, Guisse B, Greppin H & Strasser R J (1997) Biochilll Biophys Acta 1320, 95- 106

3 1 Duysens L N M ( 1979) in Chlorophyll orgall isatioll alld ellergy trallsfer ill phot05ylllhesis, (Wolstenholm G & Fitzsimens OW eds), pp 323-340, Excerpta Medica, Amsterdam